Gene Banks
Gene Banks
Gene Banks
Gene Banks
Simon H Linington and Hugh W Pritchard, Royal Botanic Gardens, London, UK
r 2001 Elsevier Inc. All rights reserved.
A gene bank is an ex situ (or offsite) collection of genetic development pressures and the not inconsiderable difficulties
material held for long-term security or for ease of access. The of managing in situ populations, the likelihood is that many
genetic material is usually in the form of live propagules or such collections will be called on in the future.
gametes though, increasingly, pure DNA is held. A bank can While in situ conservation should be seen as the primary
also be a collection of full-grown plants representing the di- goal of all conservationists, having a second line of defense is a
versity of certain species (a field gene bank) or a set of cul- prudent measure particularly if it assists in other ways. For
tures (cell, tissue, embryo, or microorganism). The widest example, by acting as a supply source, ex situ collections reduce
definition might also embrace collections of plants grown the pressure of repeated sampling of wild populations. Im-
primarily for purposes of education and horticulture in bo- portantly, through the use of such collections, access to na-
tanic gardens and animals represented in zoological gardens tional genetic resources can also be controlled. Furthermore,
or parks. because research is facilitated by such collections, knowledge
about both the conservation biology and socioeconomic value
of wild populations is improved, thereby increasing the like-
Role Within Conservation Strategies lihood of survival in situ. It is sometimes asserted that ex situ
conservation of certain species reduces political will to con-
Ex situ (or off site) conservation underwrites conservation serve the habitats in which they occur. Should this be true, it
measures in situ (on site or in the natural state) and provides would seem essential to ensure against the loss of, at the very
centers from which genetic material (germplasm) can be ac- least, flagship species within the habitats so that they can be
cessed for research, breeding, or (re-) introduction. Nearly any reintroduced. Additionally, it should be apparent that habitat
ex situ collection might be included under the loose definition complexity may limit reconstruction opportunities from ex situ
of a ‘‘gene bank.’’ As with a financial bank, deposits are made collections. In summary and as often stated, ex situ conser-
with the intention of later withdrawal. Furthermore, while in vation is complementary to that carried out in situ (see also the
the gene bank, safety and integrity will be paramount and entry for ‘‘Ex situ, in situ conservation’’).
value will be added through study. However, the emphasis The main types of animal ex situ collections are zoological
placed on withdrawal varies considerably between gene banks. gardens and parks, sperm and ova banks, and DNA banks.
For some collections, short-term utilization is the sole mo- Due to controlled breeding programs, genetic resource data-
tivation, though inevitably conservation will be served to banks also play a more prominent role in the management of
some extent. In others, long-term insurance against loss in situ such collections than is perhaps the case with those for plants
is the stronger driving force. Given the increasing risks of and particularly those in botanical garden collections. Microbe
genetic erosion (i.e., a narrowing of the gene pool leading to collections are mainly cultures though the possibility exists for
species being condemned to extinction) as a result of human fungal spore banks.
When considering plant species, the main types of ex situ A collection of medicinal plants was established in China
collections are botanic gardens specimen collections (each in 2800 B.C. Three hundred years later, the Sumarians were
normally comprising many species and few individuals per collecting vine germplasm from Asia Minor. During the Mid-
species); field gene banks (usually few species, many indi- dle Ages, the Islamic world made great strides forward in the
viduals); seed, pollen, and spore gene banks (potentially study of plants and their properties. There followed a pro-
many species, many individuals); in vitro cultures (relatively few liferation of gardens of medicinal plants during the late
species, many individuals); and DNA banks (potentially many Middle Ages and Renaissance of Europe. Acquisition and ex-
species, many individualsFthough the latter are rarely held change of crop plants gathered pace with the colonial activities
separately). The collections vary in the degree of technical input of the European powers. For instance, Columbus took wheat
required, their ability to effectively conserve inter- and intras- and other seeds to the New World (Plucknett et al., 1987).
pecific variation, their longevity, and the ease with which gene Following such relocation of crops and their subsequent
products can be obtained. Seed gene banks (in this entry ab- adaptation and diversification, the current patterns of food
breviated to ‘‘seed banks’’) nicely balance these factors and offer species use were established. Current collections of crop gen-
a very effective form of ex situ conservation for many plant etic resources reflect a strong degree of interdependence across
species. For this reason and the fact that most references to gene the world on genetic diversity born out of this era of colon-
banks relate to seed banks, this chapter will firstly consider this ization. For example, one study suggests that North America
type of gene banking and in particular the technology. now relies nearly totally on, and two-thirds of developing
countries now obtain more than 50% of their crop production
from, crops domesticated external to their region (FAO, 1996).
Seed Banks European colonization spurred on the development of
many of the world’s botanic gardens. For instance, the French
Seed Banks are an Effective Form of Ex Situ Conservation established the first tropical botanic garden at Pamplemousses
in 1735 to act as a staging post for germplasm movement and
What makes seed banks such an effective ex situ conservation to screen for use. The Royal Botanic Gardens, Kew, was es-
technique is that the methodology can be applied to a wide tablished in 1759 and in turn helped develop botanic gardens
range of species in a universal and straightforward way and elsewhere in the world (e.g., Singapore). These subsequently
that large amounts of intraspecific diversity can be conserved acted as conduits for distribution of germplasm around the
and for long periods of time without intervention. world. Many famous plant collectors date from this period of
The fecundity of most plants means that a small seed European colonial expansion and include Sir Joseph Banks,
sample can be removed from the annual seed rain with lim- Richard Spruce, and George Forest (see Plucknett et al., 1987).
ited effect either on that population’s survival or on the seed Not surprisingly, many botanic gardens now hold important
yield. One seed lot, if carefully sampled, can hold the majority collections of both endemic and exotic nondomesticated
of alleles found within that population. Moreover, if carefully species including those known to be used locally for medi-
germinated and then grown up under the same conditions, a cines and timber plus many ornamentals.
seed lot has the potential to recreate the original population. Changed agricultural practice and increased pressure on
In the meantime it can be held within a relatively small vol- the natural world during the 20th century has led to increased
ume. For instance, a million tobacco seeds, each one with the erosion of genetic diversity both in crops and wild species.
potential to grow into a genetically distinct plant, occupy Currently, the most important causes of genetic erosion are
about a fifth of a liter. considered to be replacement of local varieties, land clearance
Additionally, germinating seeds to obtain fully grown (e.g., fuel-wood collection), and overexploitation; some of the
plants is relatively simple compared with obtaining plants greatest concern relates to diversity found in the arid and
from in vitro stored material. Plants recovered from banked semiarid regions (FAO, 1996). In 1949 there were about 10
seeds can also be compared with natural populations from thousand wheat varieties in China; in the 1970s the number
which the material was harvested years before and which may had reduced to a thousand. Erosion of the genetic base of
have been subjected subsequently to environmental change crops is not new. For instance cultivated carrot color was in-
(e.g., as a result of global warming). finitely more diverse in Medieval Europe than it is now.
However, the rate of erosion has accelerated in the 20th cen-
tury with the drive toward greater genetic uniformity of crop
The Historical Context
varieties. Some highly uniform varieties have been sown across
Seed banks might be seen as an invention of the 20th century, huge areas. In 1983 a single variety was sown across two-thirds
though the exploitation of crop seeds’ ability to store in a dry of the wheat fields in Bangladesh and the following year across
state has almost certainly played a significant role in early nearly a third of all those in India. This uniformity has
human civilization. Early forms of seed storage probably in- productivity advantages within agricultural systems but is not
cluded burying the seeds in ash or sealing them in adobe huts without risk. For instance, there was a severe shortfall in winter
for the following season. By comparison, modern seed banks wheat production in the Soviet Union when a variety grown
use controlled drying facilities and deep freeze stores and aim over 15 million ha did not have sufficient cold tolerance to
for storage lives of decades, centuries, or more. Modern seed survive the winter of 1972. Less dramatic was the abandon-
banks have a lineage built on plant exploration and the de- ment in 1975 of all of the United Kingdom’s white clover
velopment of botanic gardens that dates back nearly 5 varieties due to susceptibility to the disease Sclerotinia trifo-
millennia. liorum (FAO, 1996).
Gene Banks 643
The more modern uniform varieties have been selected described higher plants. Obviously, the ex situ conservation of
from genetically heterogeneous old varieties and landraces wild plant diversity remains a major gap that needs to be filled
(primitive varieties) that they have then displaced. Therein lies within the genetic resource system.
the conundrum. For these new varieties to be developed fur- The international institutes within the Consultative Group on
ther they need to draw on the genetic diversity that they have International Agricultural Research (CGIAR) system account for
displaced. Half of the production increases of the Green 10% of the 6 million accessions. These institutes include the
Revolution were brought about by the use of plant genetic following (with number of accessions, most of which are held in
resources for plant breeding. seed banks, shown in brackets; see FAO, 1996):
The Current Status of Seed Banks for Food and Agriculture • The Centro International de Mejoramiento de Mais y Trigo
(CIMMYT) in Mexico (136,637)
Although the outstanding Russian genetic resource scientist • The International Crops Research Institute for the Semi-
Nikolai Vavilov had assembled a collection of 50,000 cereal Arid Tropics (ICRISAT) in India (110,478)
and pulses during the 1920s and 1930s, which were main- • The International Centre for Agricultural Research in the
tained by an annual grow-out, the first impetus to develop Dry Areas (ICARDA) in Syria (109,029)
modern seed banks occurred in the United States. Here it was • The International Rice Research Institute (IRRI) in the
noted in the late 1940s that less than 10% of 160,000 plant Philippines (80,646)
accessions in the national system since 1898 could still be • The Centro Internacional de Agricultura Tropical (CIAT) in
found (Plucknett et al., 1987). Following the establishment of Colombia (70,940).
U.S. regional genetic resource facilities to address this prob-
lem, the National Seed Storage Laboratory (NSSL) was estab- Just under half of the 6 million accessions are held in 12
lished at Fort Collins, Colorado, in 1958. With about 360,000 national collections. To some degree this is a function of the
accessions, it and the Institute of Crop Germplasm in Beijing early establishment of their genetic resource collections. The
with about 316,000 accessions are the world’s largest seed collections include those in Russia and the United States
banks. (noted earlier), Japan (established in 1966), Germany (1970),
A significant proportion of the accessions held in crop Canada (1970), and Brazil (1974). Of the 1308 national or
genetic resource collections (including seed banks) date from regional collections currently noted by FAO (1996), only 397
the 1960s onward when the extent of loss of diverse landraces within 75 countries are held in long- or medium-term seed
was becoming apparent. Catalyzed by the Food and Agri- banks. Medium-term storage might be assumed to be in the
culture Organization of the United Nations (FAO) and sub- order of 10 or more years.
sequently by the International Board for Plant Genetic Whilst 496 collections are within Europe and 293 are
Resources (IBPGR, now the International Plant Genetic Re- within Asia, some areas of the world have very few such fa-
sources Institute, IPGRI), a great amount of crop genetic di- cilities. FAO (1996) noted that there is only one long-term
versity was saved in the 1970s and 1980s through the store within the Caribbean and this is in Cuba. Relatively few
organization of collecting missions. Not surprisingly, conser- of the collections covered by the FAO survey are held by pri-
vation efforts have concentrated on the 30 crop species that vate companies, though many breeders hold smaller working
provide 95% of dietary energy or protein. Of these species, collections.
rice, wheat, and maize provide 26, 23, and 7%, respectively. The remits for crop banks are very varied. Some cover wide
Consequently, about 40% of the estimated 6 million acces- ranges of the crops that are important at the national or re-
sions conserved in the 1308 national and regional collections gional level, others cover very specific material. For instance,
are cereals (of which 14% are wheat). A further 15% are food facilities at Horticulture Research International in Well-
legumes. Other foods such as roots/tubers, vegetables, and esbourne (United Kingdom) and the Asian Vegetable Research
fruits each comprise less than 10% of the total. Because many and Development Centre (Taiwan) have remits for vegetable
of the species in the collections are harvested for their seeds, it germplasm.
is not surprising that most accessions (about 90%) are held While the science and technology underpinning these seed
within seed banks. The remainder are held in vitro or within banks is generally well founded (discussed later), problems
field gene banks (see later). Of the accessions for which there such as unreliable electricity supply mean that only the nine
are data, perhaps half are advanced cultivars or breeders’ lines CGIAR banks, those in 35 countries and four regional centers,
and a third are landraces and old cultivars. The tertiary (plus meet international standards as established by FAO and IPGRI
much of the secondary) gene pool within wild and weedy (1994). To some extent this problem is obviated by the high
species comprise only 15% of the accessions (FAO, 1996). The level of duplication within the world’s 6 million accessions. It
narrow range of species is indicated by the fact that forages is estimated that perhaps 1 to 2 million of these are genetically
and forest species are considered to have minimal coverage. unique.
There is no good estimate of the number of wild species held The way in which seed bank and associated genetic re-
under internationally acceptable gene bank conditions. There source work is organized both at a regional and national level
are probably more than 10,000 species, but almost certainly varies considerably. In Europe, the European Cooperative
less than 20,000 species, held. The National Plant Germplasm Programme for Crop Genetic Resources (ECP/GR) works
System in the United States accounts for some 8720 species through crop networks to coordinate effort. In Southern
(see ten Kate and Laird, 1999). This should be set against a Africa, work is coordinated through a network set up through
background of about a quarter of a million scientifically the Southern Africa Development Corporation (SADC). At a
644 Gene Banks
national level, in India, genetic resources activities are formally States and the Henry Doubleday Research Association in the
centered around the National Bureau for Plant Genetic Re- United Kingdom are examples of NGOs active at the national
sources. Within the United Kingdom, genetic resources activ- level in the conservation of traditional vegetable varieties
ities are less centralized with responsibility residing with a through their heritage seed libraries.
number of institutes and their respective ministries. Advice to
government is provided through the U.K. Plant Genetic Re-
sources Group, which draws members from all of the relevant Collection Value
organizations.
A financial value can be placed on biodiversity in one of two
ways (see FAO (1996)):
Seed Banks for Wild Species
• Direct value, or a willingness of the market to pay for it
Increasingly, botanic gardens are adopting seed banking • Indirect value, including its value as an insurance policy,
technology. Although FAO (1996) note that 150 botanic gar- the value to a product derived from the input of specific
dens (about 10% of the world total) have seed banks, rela- genetic material or the travel costs people would accept to
tively few of these have embraced international standards collect it or to visit it for tourism
(discussed later). Those that have include the ones at the Berry
Botanic Garden, Oregon (United States), the National Botanic Obviously, there are other ways of considering the value of
Garden of Belgium, the Royal Botanic Gardens, Kew (Wake- biodiversity such as its ‘‘service’’ value to biosystems and its
hurst Place), and the network of seed banks within the value as a heritage to each human generation. However, de-
Spanish botanic gardens (notably Cordoba). Most botanic bate has not surprisingly concentrated on its economic value
garden seed banks were born out of a need to more effectively and consequently the issue of access to the genetic wealth
manage the annual seed exchange. However, many now con- contained within seed banks is an increasingly controversial
centrate on the conservation of undomesticated species and subject. Behind this debate is an attempt to reconcile the fi-
interspecific variation. Additionally, several banks not in bo- nancial contribution of samples collected in one country to
tanic gardens concentrate on wild species. These include the the economy of other countries. A fundamental concern that
Threatened Flora Seed Centre established by the Department is often expressed is the degree of control over agricultural
of Conservation and Land Management in Western Australia, systems, particularly within the developing world, by multi-
the forest tree seed banks (e.g., the National Tree Seed national agrochemical/seed companies. Concern revolves
Programme in Tanzania), and the seed bank of the Uni- around the use of the following:
versidad Politecnica de Madrid. The latter bank was estab-
lished in 1966 to conserve Crucifers and species from around • Uniform F1 hybrid varieties (where the seed, if harvested
from the crop by the farmer, subsequently produces a
the Mediterranean. Together with the Royal Botanic Gardens,
heterogeneous crop of lower yield)
Kew Seed Bank that has concentrated on wild species from the
world’s drylands, it has helped pioneer the application of seed • Varieties that work best within systems when treated with
certain agrochemicals
bank technology for the conservation of wild species. Faced
with the acceleration in wild plant genetic erosion and ex- • The use of genetically modified organisms (GMOs)
tinction, the Kew Seed Bank has taken on the ambitious ob- • Plant breeders’ rights and, increasingly, patents
jective of facilitating the conservation of seed from 10% of the Set against a backdrop of a $15 billion estimated annual
higher plant species by the year 2010. Termed the Millennium turnover for the seed industry in Organization for Economic
Seed Bank Project, the work is a collaborative exercise in- Cooperation and Development (OECD) countries, the debate
volving botanical organizations from around the world. One has polarized on an ‘‘Industrial North–Developing South’’
of the aims of the Project is to maximize the efficiency of the divide. This has led to a useful reassessment of the sharing of
banking work through an active seed research and technology benefits, though inevitably there has been some over expect-
transfer program. Other than the NSSL at Fort Collins, the ation of the value of individual germplasm collections. Un-
project will have one of the largest seed research groups as- doubtedly, there are examples of large financial contributions
sociated directly with a seed bank. Certain other botanic gar- to the commercial world from plant germplasm. The value of
dens, such as that in Rio de Janeiro, also carry out seed genes for high sugar content from Lycopersicon chmielewskii
research work applicable to seed banking. have been estimated to contribute $5 to 8 million annually to
the tomato industry (see FAO, 1996). However, it is worth
bearing in mind that these are the exceptions to the rule.
Community Seed Banks
Relatively few advances in plant breeding have been spec-
A valuable role in seed banking is played by nongovernmental tacular and few have resulted from the incorporation of single
organizations (NGOs) at the community level. There is an genes. Most have occurred from the gradual improvement of
increasing use of community seed banks within the de- characteristics governed by many genes (polygenic characters).
veloping world that involve donation, storage (usually on a Of course, advanced genetic manipulation techniques may
short-term basis), and use of seeds at the local level. Examples change this picture and, indeed, may have an impact on the
include a set of community seed banks set up in North Eastern perceived value of traditional crop genetic resources.
Brazil. The Plant Genetic Research Centre in Ethiopia is an In contrast to the seed industry, the pharmaceutical sector
example of a national program integrating with a community- is estimated to be valued at $235 billion (FAO, 1996). In-
based system (Heywood, 1995). Seed Savers in the United cluded within this turnover are plant-derived drugs, many of
Gene Banks 645
which relate back to plants collected from developing coun- Particular challenges faced by seed banks include the
tries. However, large financial returns from plant-derived drugs following:
are unusual because major breakthroughs in the identification
of new bioactive compounds from plants are infrequent. It is • The acquisition of new material on terms that enable its
true that 34 out of the top 150 prescription drugs in the subsequent distribution and use, so far as possible, on a
United States have a plant-based origin. This aside, despite the common basis
estimated tens of thousands of species investigated in bio- • The development of a supply policy flexible enough to
prospecting programs, only 90 plant species have yielded 119 ensure that terms and conditions of any bilateral agree-
compounds considered to be important drugs (see ten Kate ment (e.g., with the country of origin) are met, while en-
and Laird, 1999). abling plant genetic resources to be also made available for
The debate has driven the establishment of agreements food and agriculture under any multilateral system to be
covering the access to and the sharing of benefits derived from adopted in the revised IU
plant genetic resources. Consequently, seed bank curation • The development of procedures that enable curators to
needs to accommodate not only national and international clearly identify whether a proposal to acquire or supply
legislation but compliance with increasingly strict conditions material should be dealt with on a multilateral or bilateral
attached to the material by the country or organization ori- basis. For instance, due to the single source nature of many
ginally providing the material to the seed bank. successful pharmaceutical ‘‘hits,’’ such work may fall into
The Convention on Biological Diversity (CBD) came into the latter category
force on December 29, 1993, following its opening for sig- Finally, it is important to note that the benefit of the an-
nature at the Rio Earth Summit in 1992. It has provided party nual investment in PGFRA conservation and utilization com-
governments with a framework within which access to genetic pared with its estimated total annual cost of $1 billion (FAO,
resources and the sharing of benefits arising from their util- 1996) is considered to be very high. For this reason it is es-
ization can be negotiated. Many countries have either de- sential that the future operations of seed banks are not
veloped, or are preparing national legislation to implement, threatened by financial insecurity.
access and benefit-sharing provisions of the CBD. Funda-
mental concepts that are often incorporated into national
access legislation are as follows: The Scientific Principles Underlying Seed Banking
Viable seeds of many species when maintained in a dry and
• The need to obtain prior informed consent for acquisition of
cold state are capable of being germinated many years later.
genetic resources from the appropriate national authority
This capability means that the long-term ex situ conservation
• The need to reach mutually agreed terms, which implies a
of many higher plants is a realistic possibility.
negotiation leading to a form of bilateralmaterial transfer
agreement specifying the terms and conditions under
which material is transferred Seed Storage Conditions
The science of seed storage is not a new one and dates back, at
In addition to the provisions of the CBD and imple- least, to China in the 6th century. Advances in the quantifi-
menting legislation at the national or regional level, certain cation of seed longevity under different storage conditions
seed bank collections may fall within the ambit of the Inter- were made in the second half of the 20th century through the
national Undertaking (IU) on Plant Genetic Resources for work of Harrington in the United States and Ellis and Roberts
Food and Agriculture (PGRFA). This is currently being revised in the United Kingdom (see review in Hong, Jenkins, et al.,
in order to establish a multilateral access and benefit-sharing 1998). Critical factors that determine seed longevity are the
system for PGRFA that is compatible with regimes established seed’s moisture content, temperature, and gaseous environ-
under the CBD. It also recognizes the contribution made by ment; its initial viability; and its genetic background. With
indigenous farmers, in particular, to the development of plant respect to the latter, differences between species would appear
varieties. to be much greater than those within. Genetics particularly
Seed banks, particularly the larger ones such as those influences the relationship between seed longevity and seed
within the CGIAR system (discussed previously), have re- moisture content. Most species produce seeds that can be
sponded to the emerging legal framework on access to genetic dried to low moisture contents (e.g., where less than 5% of the
resources and benefit-sharing by reviewing the terms under seeds’ fresh weight is water) without loss of viability. The seeds
which they acquire genetic resources and supply them to third of such species are termed ‘‘orthodox.’’ Furthermore, orthodox
parties. In many cases, seed banks have adopted the use of seed longevity increases in a predictable fashion as the mois-
standard material transfer agreements that entitle the pro- ture content is reduced. Within limits, there is a straight-line
viding organization (or the country of origin of the material) relationship of negative slope between longevity and seed
to a share of any benefits arising from the utilization of the moisture content (up to about 15 to 25%) when both are
material. They may also restrict the subsequent transfer and expressed on a logarithmic scale. More simply, Harrington’s
use of the material. A pioneering worldwide project by 16 rule of thumb states that a 1% reduction in moisture content
botanic gardens, which like the agricultural seed banks have roughly doubles seed longevity. Ellis et al. in 1989 found that
traditionally supplied genetic resources to a wide range of drying seeds below a certain moisture content (equating to
users, has also attempted to find broadly standard terms under that in equilibrium with air at about 11% relative humidity at
which material may be transferred to third parties. room temperature) did not increase longevity in most species
646 Gene Banks
studied. However, in a few species, a decrease in longevity was the number of wet tropical forest species producing such
noted. seeds. Seed storage behavior is usually assessed by quantifying
Temperature also has a predictable effect on seed longevity the survival of seed lots of a species after drying to different
over the range 13 1C to 90 1C (Dickie et al., 1990) such that moisture contents. A more rapid biochemical diagnosis of
there is a quadratic relationship between longevity on a potential survival of drying is clearly of interest. Most re-
logarithmic scale and temperatureFalthough an Arrhenius calcitrant seeds, including oaks and rubber, die below a value
relationship also fitted the data well. Importantly, the relative of about 40% moisture content. Because they need to be kept
effect of temperature on the seed longevity of eight species was moist (and aerated) they have short life spans, generally of a
identical. In its simplest form, Harrington’s rule of thumb few months, and are not suitable candidates for seed banking.
states that a 5 1C reduction in temperature doubles seed Other forms of genetic conservation must be employed. The
longevity. same is true for species, such as coffee, which produce
In 1980, Ellis and Roberts developed a predictive model for ‘‘intermediate’’ seeds that cannot be dried much below about
orthodox seed storage that incorporated the determinants of 10% moisture content. They also tend to be susceptible to
longevity (except gaseous environment that has minimal effect storage at cold temperatures ( 20 and 0 1C). Work on other
at the low moisture contents over which the model is ap- species, such as orchids, suggests the use of other subzero
plicable). Based on acceleration of the aging process in seed temperatures may be appropriate for the conservation of
lots by increasing their moisture content and temperature, the species sometimes included in this storage category (Pritchard
model allows extrapolation of longevity at cooler and drier et al., 1999).
conditions. For instance, rice seeds at 5% moisture content
and 20 1C (fairly typical seed bank conditions) have a Genetic Considerations
theoretical potential to survive at least 1900 years (see Hong The genetics of seed bank storage is an important issue. A
et al., 1998). Such data lend weight to occasional press reports criticism occasionally leveled at seed banks is that there is
of the germination of seeds of great antiquity, many of which selection in storage. Selection can occur during collection
cannot be confirmed due to the lack of supporting archaeo- through a biased sample (e.g., for early or late flowering
logical evidence. Nonetheless, there is, for example, evidence genotypes). It can also occur when samples are grown out
of longevity greater than 100 years at room temperature for under conditions that differ dramatically from those where the
dry wheat seeds (Steiner and Ruckenbauer, 1995). The seed seeds were harvested. There is, however, little evidence to show
storage model indicates the independence of temperature and that, compared with, say, room conditions, the seed bank
moisture content on longevity over a wide range of conditions environment does other than slow down the (normally dis-
(Ellis et al., 1995). It has been suggested that this may not tributed) times for individuals to die. In other words there is
necessarily be the case, particularly at very low moisture con- no greater risk of selection out of any individual by the con-
tents (Vertucci and Roos, 1990). The model also appears to be ditions applied than under natural conditions.
influenced by developmental stage (Hay et al., 1997). These Another criticism is that seed lots in a seed bank are pro-
deliberations do not, however, alter the current seed bank tected from evolutionary pressures that maintain fitness. The
storage recommendations (details of which are given later). extent of this problem will depend on the generation time of
Even if these longevity predictions prove to be unattainable the species involved and the severity of the selection pressures
for all species, they do imply that, for some at least, storage in (at the site from which it was collected) during the time the
seed banks may be sufficiently long to carry the current heri- seed lot was in storage. It might be argued that those species
tage of genetic material to new eras when other technologies with short generation times will quickly adapt to all but the
will be available. Predicted longevities of this magnitude also most extremely changed environments (i.e., the gene fre-
imply that the amount of regeneration of seed stocks inherent quencies in the seed lot will quickly be selected to meet the
in many gene bank programs currently may be more a func- new needs). Of course, maintaining the original genes sam-
tion of preferred working practice (e.g., regular study of col- pled from within the population may have longerterm bene-
lections), or poor application of the technology, rather than a fits of fitness for the species if pathogen attack has altered the
failure of the technology itself. natural population in the intervening period. An additional
In a study of the seed storage behavior of 6700 species, consideration is the prospect of genetic contamination of
Hong et al. (1998), found that 91% had orthodox (desiccation natural populations by chance hydridization with exotic ma-
tolerant) seed. The remainder had either desiccation suscep- terial including genetically modified organisms. This concern
tible or ‘‘recalcitrant’’ seed or had characteristics intermediate is also partially alleviated by the existence of long-term stored
between the recalcitrant and orthodox types. Also certain germplasm in its original state.
genera (e.g., Acer) contain both orthodox and recalcitrant
species. There is little evidence, however, that individual spe-
cies produce both orthodox and recalcitrant mature seeds. It Seed Bank Management
should, though, be noted that orthodox seed passes through a
Procedure
desiccation susceptible phase during development. The true
The basic elements of the seed banking procedure (more or
percentage of species with orthodox seed within the plant
less in order) are as follows:
kingdom is probably significantly less than 91% as the sample
was biased toward species in trade. Although recalcitrant seed • Collection planning and permission seeking
behavior is loosely associated with long-lived perennial spe- • Seed (and pressed specimen) collecting and field data
cies producing large fleshy fruits, there is great uncertainty of recording
Gene Banks 647
• Shipment of the seeds could diminish both the unique genetic makeup and reduce
• Creation of a data record about the accession the longevity (through lower initial seed quality) of a seed lot.
• Seed cleaning (sometimes preceded by initial drying and
sometimes accompanied by X-ray analysis and quantity
determination) Seed Cleaning
• Main drying Seed lots harvested from some populations of wild species in
• Seed moisture determination particular contain large numbers of ‘‘empty’’ (or aborted) and
• Initial germination test (sometimes left until after banking) insect-infested seeds. They may be outwardly similar to the
• Packaging and banking and security duplication ‘‘competent’’ seed and often are thus not removed during the
• Characterization (including verification of identity in the cleaning process. Their presence can be detected by X-ray an-
case of wild species) and evaluation (where appropriate) alysis or a simple cut-test applied to a subsample. Once de-
• Distribution of stocks to users (through time) tected, such incompetent seeds need to be allowed for when
• Germination retests (through time) distributing seed to users and when carrying out germination
• Regeneration/multiplication (as required) tests. Although the insects may not be removed at this stage,
nearly all adult and most larval forms of insects are killed by
Although many of the well-established seed banks broadly
standard seed bank practice. However, insect eggs along with
follow these procedures, most modify them to meet their own
fungal and viral contaminants can survive seed banking. Thus,
specific needs. Some banks carry out very little field collecting
appropriate quarantine procedures need to be in place when
and some multiply the seed sample on arrival. Costs for each
the seed is removed from store.
stage vary considerably between seed banks. The RBG Kew
Seed Bank, which has an international collecting program,
estimated, in 1997, that the ratio of costs between collecting, Seed Bank Storage Standards
processing, and annual maintenance for a collection was in No two banks are the same. Traditionally, seed banks have
the order of 100:50:1. been classified into the following categories:
Seed Collecting
• Base collections that are for the long-term storage of seed
lots and from which seeds are not normally sent to users
Set against the background of the CBD and the IU, collecting
(though this is not always the case).
should only be carried out with the permission of the national
and local authorities. Foreign collectors should work col-
• Active collections from which seeds are made available to
users. Often such stores are maintained under less optimal
laboratively with local scientists and clear agreements on
storage conditions compared to those holding base col-
benefit sharing should be in place. One immediately tangible
lections though this need not be the case.FAO/IPGRI
benefit is the sharing of collections and the information re-
(1994) recommends that storage lives of 10 to 20 years
lating to them. Other international regulations need to be
might be appropriate.
adhered to. These include the Convention on International
Trade in Endangered Species (CITES) and national quarantine Base and active collections can occur at the same or dif-
laws. ferent genetic resources centers or, as is the case in the SADC
Seed collecting methodology and genetic resource explor- genetic resources network, the active facilities in different
ation have been thoroughly covered by Guarino et al. (1995). countries are linked to a regional base facility. In theory, the
In most instances, random and even sampling of wild plant or collections held within the active store need occasionally to be
crop populations is recommended, including careful note refreshed by the same samples held in the base store. Because
taking of the sample method (in often less than perfect con- active stores are likely to more frequently regenerate (dis-
ditions). Objective data recording is essential as is accurate cussed later) their samples, selection and genetic drift could
recording of location. This latter aspect is now facilitated by cause a genetic divergence from the same samples held in the
the use of Global Positioning System receivers that help fix base store.
latitude, longitude, and even altitude using satellites. The FAO/IPGRI standards (1994) for base collection stor-
Harvesting seed that is close to maturity and keeping the age requires fresh seed to be dried within the range of 3 to 7%
seed alive in the field should be paramount. Also it is essential moisture content, packaged in a moistureproof way, and
that harvested seed is returned to the seed bank facility as placed at subzero (and preferably 18 1C) temperatures. Al-
securely and rapidly as possible. Prestorage conditions influ- though many of the world’s seed banks aspire to such con-
ence seed viability at the start of storage. Because the longevity ditions, as has been noted, relatively few are able to emulate
of a species stored under specific seed bank conditions is them. Nonetheless, the robust nature of seed bank technology
substantially fixed, the only other key longevity factor that can means that substantial longevities are achievable even with
be varied is initial seed storage viability. Delay in drying the partial fulfillment of the standards that have been discussed
seed properly or unduly harsh cleaning methods can severely and using a low technology approach.
reduce initial viability; in turn, this can dramatically reduce The standards set out in 1994 are important as they set the
both seed longevity and seed bank efficiency. Where space is benchmark for standard operating procedures by seed banks.
not limiting within a cold store and the longer seed lots can be Consolidation of these standards and increased use of a
kept alive, the less the unit fuel cost per species. common format for data would facilitate exchange of infor-
It is worth noting that seed lots of the same species should mation between facilities. Such unification of data standards
not be mixed either within or between years. Such action could ultimately embrace all specimen banks including those
648 Gene Banks
specializing in the storage of samples for environmental cooled, and then returned. In effect, this is a closed system
monitoring. though most facilities allow for some fresh air intake. Ducting
the air to and from the dryer on opposite sides of the chamber
Seed Drying encourages air movement. Conditions within the chamber are
Drying is perhaps the key to success in seed banking of those necessary to achieve the desired moisture contents at
orthodox-seeded species. Not only is storage prolonged in equilibrium for long-term storage. Such conditions may be 10
orthodox-seeded species but germination is prevented, the to 15% relative humidity at 10 to 25 1C, and equilibrium is
risks of insect and mite attack are reduced, and the seeds are usually achieved in about one month.
protected from freezing damage. FAO (1996) found good
drying facilities to be a limiting factor in a number of seed Packaging
banks. Drying involves the manipulation of the water poten- Maximizing orthodox seed longevity depends on drying and
tial gradient between the inside and the outside of the seed. To freezing. Cooling ambient air increases its relative humidity
all intents, the water potential is the difference between the and hence its water potential. Placing the dried seeds directly
activity of water molecules in any system compared with pure inside a refrigerator will cause the seeds to absorb moisture.
water. When the water potential of the air is lower (more Therefore seed banks need to carefully package the dried
negative) than that of the seed, there is a net movement of samples. In 1996, IPGRI reviewed the types of container used
water out of the seed and drying occurs. As the water potential in conventional long-term seed stores. Types of container used
gradient reduces, drying slows down. When no gradient exists, by banks include laminated aluminum foil bags, sealed steel
the seed moisture content is said to be in equilibrium with the cans, sealed glass tubes, screw-top glass bottles, and levertop
surrounding air conditions. Consequently, factors that influ- fruit preserving jars. All containers have some limitations but,
ence the water potential of the air surrounding the seed are carefully managed, risks of moisture ingress can be minimized
important to drying. Lowering the relative humidity or in- especially if some monitoring system is in place. The type of
creasing the temperature of the air lowers its water potential. container chosen will partly depend on the frequency of access
By keeping air moving over the seed, low humidity can be required. The main concern with all is that the seal through
maintained by preventing moist air from accumulating ingress of moisture through the fabric of the material over
around the surface of the seed. Seed size and seed depth affect long periods is a risk where foil is punctured or cans have poor
rate of drying. Migration of moisture from the center of a large seams. Glass offers the advantage that the contents are visible
seed to the outside will take longer than from the center but is of course at risk from breakage. A number of banks
of a small seed. The same principles apply to large and small double-pack for added security and others add a desiccant.
seed sacks. Shape, affecting surface to volume ratios, and seed A number of facilities dry the air in the cold store and store the
structure will also have an effect on drying rate as can seed collections in paper or cloth bags. Consequently, these facil-
maturity. ities must have adequate generator backup. This is less of a
It is worth noting that at any particular water potential, the problem where seeds are held in moisture-proof containers
seed moisture content will depend on its chemical com- and there is a loss of electrical power. In such circumstances
position. Seeds with a higher oil content will have a lower there is little evidence to suggest that more than a few days
moisture content. storage life is lost per disruption.
Most seed banks are located where the ambient relative
humidity is not sufficiently low to allow the seeds to dry to the Storage Temperature
moisture content levels set out in the previous standards. High Many long-term seed banks store seed under deep freeze
temperature gives rapid drying. However, although shade and conditions using either purpose-built prefabricated cold
oven drying can be used for drying, there are seed aging rooms or domestic deep freezers. To reduce staff time at sub-
dangers of placing wet seeds at high temperatures and of zero temperatures, a few seed bank cold rooms, such as one at
leaving dried seeds too long under such regimes. Similarly, sun the National Institute of Agrobiological Resources (NIAR),
drying has risks associated with radiant heat gain. A degree of Japan, have mechanized banking/retrieval systems. The use of
caution needs to be exercised if these methods are used. such systems have implications to energy consumption by the
Consequently, seeds are usually placed in a drying environ- bank.
ment where the relative humidity has been artificially lowered. Use of permafrost has been considered for long-term du-
Most often, this is achieved by sorption or occasionally by plicate storage of seed in places such as Svalbard. Although the
refrigeration drying systems. At their simplest, sorption sys- dependence on electricity is cut, such stores are usually unable
tems can consist of a closed container into which the wet seeds to match the lowering of temperatures possible in con-
are placed and the air dried by a quantity of silica gel or dried ventional base storage conditions. In 1997 the Japanese-based
rice. Such methods require a degree of experimentation to Biological and Environmental Specimen Time (BEST) Capsule
achieve the desired results. Greater control over the extent of 2001 Project discussed the possibilities of long-term storage of
drying can be achieved by use of a suitable saturated salt so- flagship samples under Antarctic ice at 58 1C (which inci-
lution (e.g., that of lithium chloride) that will maintain a set dentally is not sufficiently low for animal tissue preservation)
relative humidity (about 11% at room temperature) within a or even on the dark side of the moon at 230 1C.
closed container. However, many of the larger seed banks have More usually, seed storage at ultra-low temperatures
now adopted controlled drying rooms that allow for large (cryopreservation) is achieved using liquid nitrogen. Seed
samples to be dried in thin layers. Air from the chamber is samples are normally held in polypropylene (or similar)
dried using dryers containing lithium chloride or silica gel, screw-cap containers placed in the vapor phase above liquid
Gene Banks 649
nitrogen (about 160 1C). Preferably samples of dry seed to take place, sufficient seeds should remain in the collection
should be cooled and rewarmed at a relatively slow rate to make several attempts at growing out the collection. Be-
(about 10 1C min 1) to reduce problems of rapid thermal cause base collections may have a projected storage life of as
contraction and expansion that can cause physical injury to much as 200 years, collection size needs to be large. The
the seeds, such as cracking of the embryo tissue. The largest international standards (discussed earlier) recommend at least
cryogenic seed bank is operated by the National Seed Storage 1000 seeds per seed lot.
Laboratory (United States) where there are more than 37,000 Where seed banks handle seed with unknown storage be-
accessions stored over liquid nitrogen. More than 11,000 of havior, the initial germination could be delayed until after
these are also stored under conventional seed bank conditions drying and banking. Survival will indicate orthodox charac-
at 18 1C. The National Bureau for Plant Genetic Resources teristics. Few banks will have sufficient staff time to do more
in New Delhi also conserves seeds under conventional and than brief tests prior to this on suspected recalcitrant or
cryogenic storage conditions. Moreover, seeds of a significant intermediate seeded species.
number of North American and Australian wild species have
been tested successfully for their tolerance of cryopreservation
Duplication
(Pence, 1991; Touchell and Dixon, 1993). It is worth bearing
One of the main advantages of seed banks is that they cen-
in mind when considering setting up a cryogenic facility for
tralize collections of genetic material making them more easily
seed material that such low temperatures are not necessary to
accessed and studied. Indeed, some seed banks might be seen
achieve practical periods of long-term storage (Pritchard,
as some of the world’s greatest plant diversity hot spots with
1995), that the setup costs are relatively expensive, and the
more individuals and, in some banks, more species per square
storage volume is less efficient than a conventional seed bank.
meter than anywhere else on the planet. This centralization
However, these additional costs may be acceptable when cre-
poses a risk to all but the most carefully located and con-
ating the ultimate ‘‘base collection’’ for material that is in short
structed facilities. Potential catastrophic loss, which of course
supply, is inherently short-lived, or is of particular commercial
threatens plants conserved both ex situ and in situ, mean that
value (unique genotypes).
duplication of collections and their associated data is an im-
portant element of seed bank safety. The FAO report (1996)
Monitoring Seed Lot Viability indicates that the level of security duplication of plant genetic
Perhaps one of the most important parameters of seed bank
resources for food and agriculture still needs to be improved
effectiveness is the result of germination monitoring. Ger-
and is at best uncertain.
mination is the preferred test for seed lot viability. Providing
such information to those using the seed is helpful. Add-
itionally, other viability tests such as vital staining using tet-
Characterization and Evaluation
Characterization can vary from accurate naming of the species
razolium solution have a greater element of subjectivity about
or subspecies represented by the collection through to more
them. This staining test is, however, sometimes used to help
detailed recording of characters governed by genes that are
distinguish between dead and dormant seeds among those
little modified by environmental factors (major genes). Such
that did not germinate under a given test regime. Two prob-
information, published in the form of descriptor lists, is of
lems relate to the germination monitoring of seed bank ac-
great value to plant breeders wishing to narrow their choice
cessions. First, because seeds are tested soon after arrival at the
of material from, often, vast collections. Similarly, the concept
bank and then at regular intervals (often every 5 to 10 years)
of core collections has been established to facilitate use by
during their storage life, the tests need to be repeatable and
breeders. A core collection genetically represents a limited set
operator independent. Second, in order to recover as many
of accessions of a crop gene pool with the minimum of
genotypes as possible represented within a seed lot, it is ne-
repetition.
cessary to break seed dormancy. This can be a particular
Increasingly, characterization is taking the form of more
problem in seed of wild species and where the seed is freshly
detailed molecular techniques such as screening by Amplified
harvested. Key techniques include scarification of hard seed
Fragment Length Polymorphism. By contrast to character-
coats to facilitate water or oxygen permeation, imbibed chil-
ization, evaluation records data on traits such as yield that are
ling at 5 to 10 1C, and later incubation at diurnal alternating
strongly influenced by the environment in which the plants
temperatures with fluorescent light (i.e., rich in red light)
are grown. Such data are thus site and year specific and are
provided only in the higher temperature phase. Tests have to
perhaps of less use to breeders.
be seed lot specific as most seed dormancy is not strongly
genetically inherited and the form it takes depends on the
conditions under which the seed matured. Once determined, Distribution to Users
the same treatments can be used during the monitoring of that A very important element of seed bank work is to make the
seed lot through time. seed available wherever possible. In evidence of the scale of
Seed banks use a variety of media for germinating the such dispatch, ten Kate and Laird (1999) quote an annual
seeds, such as filter paper and sterilized sand. However, 1% distribution of nearly 120,000 samples by the U.S. National
(w/v) plain water agar is increasingly popular especially as it Plant Germplasm System of which 65% are sent abroad, many
reduces the risk of imbibition damage to dry seeds. requested by reference to the Germplasm Resources Infor-
Obviously through time these monitoring tests can con- mation Network (GRIN) available on the Internet. Further-
sume a significant proportion of each seed lot. Ideally, when more, the usage rate of crop banks by plant breeders is
the seed viability has fallen to a level where regeneration needs probably less than the rate of request from banks holding
650 Gene Banks
broader plant diversity collections where uses include a Other Types of Gene Bank
wide array of pure and applied research in addition to field
trials. During 1994–1996, there was a 50% request rate for This section provides summaries of the current status of
seeds offered through an extensive list offered by the Kew nonseed gene banks, starting with dry propagules (pollen and
Seed Bank. spores), which can be stored under conditions similar to those
Several elements need to be considered concerning the used for seeds, and covering normally hydrated tissues that
distribution of seed to users. The recipient should be provided can also be preserved under a different set of controlled con-
with accurate information about the collection and how to ditions. Finally, field gene banks are covered and the role of
germinate the seeds. It should also be remembered that certain botanical and zoological gardens is briefly mentioned.
species require a symbiont for effective growth (e.g., legumes
and Rhizobium), and that the user may need to draw on
germplasm for both plant and symbiont. The seed sample Pollen
must be dispatched having fulfilled all necessary plant health There are many practical reasons for storing pollen: to support
and, where appropriate, CITES requirements. Finally, to meet work on allergenic responses, plant hybridization, and fer-
the needs of the CBD and to clarify the conditions under tility; haploid plant production; and genetic transformation
which the material can be used, all germplasm samples are systems with isolated pollen (or gametes). Optimal storage
increasingly dispatched under material supply agreements. conditions for pollen are similar to those for seeds (i.e., at low
moisture content and subzero temperatures). In addition,
Regeneration there is some evidence (e.g., in maize and Impatiens) that
Seed bank accessions are grown out for the purposes of re- longevity in dry storage is enhanced in anoxic atmospheres.
generation of seed stock (either when seed numbers are low or Moreover, pollen has also been stored in a vacuum-dried state.
when viability has reduced), for characterization and for The ease of storage though relates in part to the cellular and
evaluation. Many banks have a regeneration standard below physiological nature of the pollen. Bicellular pollen, as found
which the germination of a seed lot should not fall. This is in Liliaceae, Orchidaceae, Solanaceae, and Rosaceae, generally
usually set at 85%. This high value limits the risk of accu- tolerates desiccation to about 10% moisture content and is
mulated genetic damage that is associated with seed aging. relatively long-lived. By comparison, tricellular pollen, as
Even though falling levels of seed germination are correlated found in Graminae and Compositae, is relatively short-lived
with falling levels of field establishment, many banks have and is much more sensitive to desiccation. Most work on the
adopted lower standards. This may in part be due to the long-term storage of pollen has focused on fruit tree or for-
backlog of regeneration work that in some national facilities estry species, for which 10 years storage at conventional gene
highlighted by FAO (1996) is nearly 100% of the collection. bank temperatures ( 20 1C) is easily attainable. Pollen of at
By collecting high-quality seed lots in good quantity, other least 30 species are known to survive liquid nitrogen tem-
banks have reduced the necessity for regeneration that can be peratures. Although pollen banking is evidently possible for
time and labor consuming and that can have adverse effects many species, there does not appear to be any large-scale gene
on the genetics of the collection. Samples regenerated under bank operation using such material.
conditions different from where they originated can experi-
ence selection. If too small a sample is regenerated, genetic
drift may occur in which rarer alleles are lost through chance. Spores
Under some circumstances, recollection, if possible, may the Spores of many species of both pteridophytes and bryophytes
more desirable option. are stable for months or years when dried, and this time can
be extended with storage at cold or freezing temperatures. The
Seed Bank Design longevity of short-lived (chlorophyllous) spores of some spe-
Having considered the aspects of seed bank management, a cies can be extended significantly by drying and freezing in
brief consideration of seed bank design is appropriate (also liquid nitrogen (Pence, 2000). Although there are data on
see Cromarty et al., 1985). The location of the bank is im- fungal spore storage (e.g., work by Hong et al. in 1998), the
portant from political, practical, and security aspects. Potential majority of fungal germplasm appears to be conserved in
risks have to be considered be they earthquake, flooding, or culture or through cryopreservation of hyphae (see later).
radiation fallout. Some facilities are placed underground such
as the seed bank at Krasnodar in Russia and the Millennium
Somatic and Zygotic Embryos of Plants with Nonbankable
Seed Bank in the United Kingdom. Others such as the NSSL
Seeds
are located on the first floor to limit possible impact from
structures above resultant from seismic activity. The size of Nonbankable seeds can nonetheless be stored using alter-
most banks should be dictated by peak annual intake (seed native approaches. Usually, these revolve around the use of
drying and cleaning facilities), projected capacity before a re- rapid, partial desiccation of embryos or embryonic axes to
build is practical (seed storage), and annual collection main- about 20% moisture content and subsequent transferal to li-
tenance (germination, field, and greenhouse facilities). Cold quid nitrogen temperature or the use of other subzero storage
storage facilities vary from a few domestic deep-freezers up to temperatures. Recovery levels may be improved by pretreat-
large rooms such as one of 140 m2 (with capacity for 150,000 ment of embryos with cryoprotectants, encapsulation of the
samples) at NIAR in Japan. material in alginate beads, or careful manipulation of the
Gene Banks 651
in vitro recovery environment. The embryos of more than 50 (usually meristem or nodal cutting) is frequently transferred
species have been successfully cryopreserved. Other parts of (subcultured) to fresh nutrient medium with the risk of mi-
the plant can also be used to establish ex situ gene banks for crobial contamination, or loss through human error. To retard
species with nonbankable seeds, as described in the following growth and hence extend the subculture interval, temperature
section. and light intensity are reduced. For example, 0 to 5 1C and
1000 Lux are generally used for cold tolerant species, and 15 to
22 1C and reduced light intensity for tropical species. Alter-
Vegetative Parts of Plants
natively, growth can be slowed down by the addition to the
Vegetative tissues of both pteridophytes and bryophytes can be medium of chemicals to induce mild osmotic stress (e.g.,
banked for germplasm preservation (Pence, 2000). Gameto- mannitol) or hormonal retardants (paclobutrazol, abscisic
phytes of many bryophyte species are naturally adapted to acid). Also, maintenance of tissue under reduced oxygen (e.g.,
desiccation stress and can be cryopreserved after sufficient under mineral oil or liquid medium) slows growth. Under the
drying. Gametophytes of pteridophytes and some desiccation- appropriate conditions subculture intervals can be extended to
intolerant bryophytes can also be frozen when provided with one year or longer. The slow growth technique is now rou-
some cryoprotection, such as encapsulation in alginate beads tinely used for the medium-term conservation of a number of
followed by dehydration or the use of abscisic acid and the species such as banana, potato, yam, cassava, and strawberry.
amino acid, proline, as a pre-treatment. Shoot tip freezing of Although in vitro culture without cryopreservation poses
sporophytes of pteridophytes has also been demonstrated. It is considerable threat of genetic drift, the propagation of plant
estimated that fewer than 200 taxa of bryophytes and pter- material in an aseptic environment ensures the production of
idophytes combined are currently banked worldwide using disease-free stock material, which is readily accessible inter-
vegetative tissues, primarily at the Cincinnati Zoo and Bo- nationally because it satisfies most country quarantine re-
tanical Garden and the University of Kansas in the United quirements. Undoubtedly, in vitro culture is a valuable
States, but there is significant potential for increasing this complementary approach to field conservation and is par-
number. ticularly useful when applied to species that are predominately
The National Seed Storage Laboratory of the U.S. Depart- propagated vegetatively (banana, potato, and pear), produce
ment of Agriculture (USDA) also cryopreserves about 1700 non-bankable or highly heterozygous seeds, and have a par-
apple lines using dormant scion sections, which are retrieved ticular gene combination (i.e., elite genotypes; see Ashmore,
by grafting (i.e., no culture of meristems). The lines are mainly 1997). The impact of these positive features of the technique is
from Malus x domestica, plus 10 to 15 other apple species. such that FAO estimates that 37,600 accessions of plant ma-
Some pear and cherry species (Towill and Forsline, 1999) are terial (vegetative and embryos) are conserved in vitro (in-
also banked in this way. cluding cryopreservation) worldwide.
Apical shoot tips and other meristems/buds are the most
popular vegetative materials for cryopreservation. Lines from
about 50 species are now routinely cryopreserved. Initial
Animal Germplasm Samples
studies used shoot tips from cold hardy, temperate zone spe-
cies (apple, pear), but cryopreservation methods have been A majority of ex situ animal germplasm is maintained in
extended to tropical zone species (banana, pineapple). Suc- zoological gardens and institutes around the world. The NIAR
cessful cryopreservation depends on defining the physiological in Japan holds 621 accessions of animal germplasm, including
adaptation of the stock plant, the explant size and type, and its silk worms in the living state. More than 100 of these acces-
water content, the steps in the preservation process (cryo- sions, mostly sperm, are cryopreserved (for a general meth-
protectant concentrations and rates of addition/removal; odology, see the discussion presented later). Similarly, the
cooling/warming rates), and the recovery system. Two-step main gene bank methodology for ova is cryopreservation.
cooling procedures have been useful for some species, but At present, however, procedures for sperm and ova preser-
vitrification procedures (solution-based systems and en- vation are not well developed for wild species, even though a
capsulation/dehydration systems) are more favored because of number of reported successes with artificial insemination and
the technical simplicity (Sakai, 1993). All methods are de- frozen semen can be found in the literature, especially for
signed to reduce ice crystal growth in the specimen. Other ungulates such as deer and antelopes. The concept of gamete
vegetative material that has been cryopreserved using similar rescue from tissues has considerable value for spermatozoa,
methodological approaches include cell suspensions and cal- where epididymal spermatozoa are readily obtainable post
lus (more than 40 species), protoplasts (more than 10 spe- mortem and can be frozen using glycerol as a reasonably
cies), and root cultures (5 species). standard cryoprotectant. Oocyte cryopreservation has only
It should be noted that cryopreservation of vegetative been achieved in the hamster, rat, rabbit, and cow and is
germplasm overcomes the problem of genetic instability dur- therefore not a practical proposition at present. Interest in
ing storage as all cellular divisions and metabolic processes are freezing ovarian tissue, and then culturing follicles and oocytes
stopped. In contrast, two other methods of in vitro preser- by various methods after thawing, has recently been resur-
vation, normal and slow growth techniques, run the risk of rected and progress has included the birth of a lamb
genetic changes (somaclonal variation) in the conserved originating from ovarian tissue autotransplanted into the
germplasm that may result in the loss of distinct genotypes. donor-recipient after freezing and thawing. In another recent
Species are stored under normal growth conditions (e.g., study, isolated rat spematogenic cells were transferred to a
Coffea at 27 1C) for short-term purposes only. The explant mouse testis, where they displayed the ability to develop into
652 Gene Banks
Agricultural Research Service Culture Peoria, IL (US) 78,010 Algae, bacteria, fungi, yeasts, actiomycetes
Collection, USDA
American Type Culture Collection (ATCC) Rockville, MA (US) 53,615 Algae, bacteria, fungi, yeasts, protozoa, cell
lines, hybridomas, viruses, vectors,
plasmids, phages
CABI Bioscience UK Centre (formerly (IMI) Egham, UK 21,000 Fungi, bacteria, yeasts
Centraalbureau voor Schimmelcultures Baarn, The Netherlands 41,300 Fungi, yeasts, lichens, plasmids
(CBS)
Culture Collection, University of Goteborg Goteborg, Sweden 28,100 Bacteria, fungi, yeasts
(CCUG)
spermatozoa. The testicular cells were frozen-thawed prior to Miyazaki, 1999). Their on-line database lists species held and
transplantation and development, and thus there may be some the expertise and services provided by the collections along
merit in exploring the cryopreservation of testicular cell sus- with contact addresses and links to collection websites. There
pensions as an alternative or adjunct to the preservation of are about 11,500 species held. Over 25% of the strains are held
spermatozoa. Cell suspension from genetically important ani- by 5 of the 497 collections (see Table 1).
mals could be used to populate the testes of common species,
thus permitting the eventual harvesting of spermatozoa. It
should be stressed that this technique is still only in its infancy. DNA Banks
Unlike animal species in which reproductive cells and tis-
DNA banks have been established in several places worldwide
sues are stored to conserve the gene pool, in humans they are
(Adams, 1997), the largest (more than 140,000 clones) being
stored for the use of the couple/woman electing to have them
for plant and animal material at the NIAR in Japan. This
cryopreserved. Some embryos are subsequently donated for the
collection is mainly constituted of rice clones (about 36,000
treatment of others. Storage is limited by law in some countries.
comprising Random Fragment Length Polymorphisms, cDNA,
The storage of embryos is common practice in in vitro fer-
YAC) and pig clones (about 106,000 comprising cosmids,
tilization (IVF) clinics worldwide, with about 70 offering the
BAC, cDNA). The most diverse DNA bank for plants is at the
service in the United Kingdom alone. Oocyte cryopreservation
Royal Botanic Gardens (RBG) Kew, which currently holds
is poorly developed and few clinics store them other than for
more than 10,000 DNA samples from a wide range of species.
research. An increasing number are offering tissue storage.
Standards of quality of preservation differ between banks and
Despite sporadic reports of cryopreservation using none-
some, such as those at the Missouri Botanical Gardens (St.
quilibrium rate cooling, the vast majority of clinics prefer
Louis, Missouri, in the United States) and several zoos, store
conventional slow cooling rate procedures with the samples
only frozen tissue. Others extract DNA and purify it to varying
loaded into ‘‘straws’’ (embryos and oocytes) or plastic vials
degrees. Purposes for these banks differ as well, with some
(ovary). The first step involves the addition of the cryo-
established to hold samples of a particular country or region
protectant (about 1.5 M; 1–2 propanediol 7 sucrose (pro-
with the intent of using these in conservation genetic studies,
nucleate and early cleavage stage embryos), dimethyl sulfoxide
whereas others focus more on taxonomic and systematic
(4–8 cell embryos, and ovary) or glycerol (blastocyst stage
studies, such as that at RBG Kew. Most banks are prepared to
embryos)). Ice formation is then induced at 5 to 7 1C and
consider sharing aliquots of DNA or small samples of tissue
the samples are cooled further at rates of about 0.3 to 0.5 1C
with researchers at other institutions.
per minute to various subzero temperatures before storage in
liquid nitrogen. After warming at appropriate rates (of about
20 to more than 300 1C per minute), the samples are returned Field Gene Banks
to isotonic conditions stepwise, with or without the addition
of sucrose to the diluent. Field gene banks are ex situ collections of mainly agricultural
or forest species. They should be contrasted with what might
be termed ‘‘farm gene banks’’ where crop germplasm is, in
Microorganisms effect, conserved in situ by the farmer. Field gene banks nor-
mally comprise considerably more individuals per accession
The most widely applicable preservation method for the pre-
than is the case in botanic gardens. Their particular use is for
servation of microorganisms that retains viability and stability
the conservation and utilization of species with the following
is cryopreservation. However, for convenience and ease of
traits:
transport freeze-drying is preferred for most bacteria, viruses,
and sporulating fungi (see Hunter-Cevera and Belt, 1996). • Have nonbankable seeds
There are currently 497 collections from 60 countries • Have long life cycles where growing up material for regular
maintaining microorganisms registered with the World Data study from a seed collection is impractical
Centre for Microorganisms in Japan (see Sugawara and • Are normally vegetatively propagated
Gene Banks 653
Essentially, they are not a new idea as the Kayapo people of References
Brazil maintain germplasm collections of tuberous plants in
hillside gardens protected from flood (see Plucknett et al., Adams RP (1997) Conservation of DNA: DNA banking. In: Callow JA, Ford-Lloyd
1987). Most countries have at least one field gene bank and BV, and Newbury HJ (eds.) Biotechnology and Plant Genetic Resources:
FAO (1996) estimate some 527,000 accessions are conserved Conservation and Use, pp. 163–174. Wallingford: CAB International.
Ashmore SE (1997) Status Report on the Development and Application of in Vitro
in this way worldwide. Examples of field gene banks include Techniques for the Conservation and Use of Plant Genetic Resources. Rome:
the National Fruit Collection in the United Kindgom, one for IPGRI.
cassava at CIAT, Colombia, one for sugarcane at the Centro Cromarty AS, Ellis RH, and Roberts EH (1985) The Design of Seed Storage
Nacional de Pesquisa de Recursos Geneticos e Biotecnologia Facilities for Genetic Conservation. Rome: IBPGR.
Dickie JB, Ellis RH, Kraak HL, Ryder K, and Tompsett PB (1990) Temperature and
(CENARGEN), Brazil, and the potato collection at the Centro
seed storage longevity. Annals of Botany 65: 197–204.
Internacional de la Papa (CIP) in Peru. Such facilities are Ellis RH, Hong TD, and Roberts EH (1995) Survival and vigour of lettuce (Lactuca
considered by FAO to be particularly important in small island sativa L.) and sunflower (Helianthus annuus L.) seeds stored at low and very-
developing states. It should also be noted that the Nordic low moisture contents. Annals of Botany 76: 521–534.
Gene Bank, which operates on a regional basis, unlike many FAO (Food and Agriculture Organization of the United Nations) (1996) The State of
the World’s Plant Genetic Resources for Food and Agriculture. Rome.
seed banks, includes within its remit the in situ conservation of FAO/IPGRI (1994). Genebank Standards. Rome.
wild crop relatives. Guarino L, Ramantha Rao V, and Reid R (eds.) (1995) Collecting Plant Diversity.
While they offer the opportunity for characterization and Technical Guidelines. CAB International.
evaluation, such collections are labor intensive and are sus- Hay FR, Probert RJ, and Smith RD (1997) The effect of maturity on the moisture
relations of seed longevity in foxglove, (Digitalis purpurea L.). Seed Science
ceptible to catastrophic events. For example, a field collection
Research 7, 341–349.
of yams was lost in St. Lucia during 1994 as the result of Heywood VH (ed.) (1995) Global Biodiversity Assessment. Cambridge: Published
cyclone damage. This is one of the reasons that a number of for the United Nations Environment Programme by Cambridge University Press.
field gene bank collections are now backed up in vitro. Hong TD, Jenkins NE, Ellis RH, and Moore D (1998) Limits to the negative
logarithmic relationship between moisture content and longevity in conidia of
Metarhizium flavoviride. Annals of Botany 81: 625–630.
Botanical and Zoological Gardens Hong TD, Linington S, and Ellis RH (1998) Compendium of Information on Seed
Storage Behaviour. 2 volumes. Royal Botanic Gardens, Kew, United Kingdom in
Botanical and zoological gardens may be seen as types of gene collaboration with the University of Reading and IPGRI.
bank with relatively few individuals per accession. While zoos Hunter-Cevera JC and Belt A (eds.) (1996) Preservation and Maintenance of
Cultures used in Biotechnology and Industry. San Diego, CA: Academic Press.
have embraced careful breeding programs that help maximize
ten Kate K and Laird SA (1999) The Commercial Use of Biodiversity: Access to
the genetic value of the limited collections across the world, Genetic Resources and Benefit-Sharing. Oxford, UK: Earthscan Publications.
this is much less true of collections in botanical gardens. Here Pence VC (1991) Cryopreservation of seeds of Ohio native plants and related
species may be represented within the ‘‘botanical gardens species. Seed Science and Technology 19: 235–251.
flora’ by a single genotype such is the clonal exchange of Pence VC (2000) Ex situ conservation methods for bryophytes and pteridophytes.
In: Havens K, Guerrant E, and Maunder M (eds.) Saving the Pieces: The Value,
material. Limits and Practice of Offsite Plant Conservation. In preparation.
Plucknett DL, Smith NJH, Williams JT, and Anishetty NM (1987) Gene Banks and
the World’s Food. Princeton, NJ: Princeton University Press.
Acknowledgements Pritchard HW (1995) Seed cryopresevation. In: McLellan MR and Day JG (eds.)
Methods in Molecular Biology Volume 38: Cryopreservation and Freeze-Drying
Protocols, pp. 133–144. Totowa, NJ: Humana Press.
The authors would particularly wish to acknowledge the fol- Pritchard HW, Poynter ALC, and Seaton PT (1999) Interspecific variation in orchid
lowing for their assistance in providing information: Mr. M. seed longevity in relation to ultra-dry storage and cryopreservation. Lindleyana
Ambrose (John Innes Centre, United Kingdom); Dr. D. Astley 14(2): 92–101.
(HRI, United Kingdom); Dr. M. Fay, Mr. R. Smith and Mr. M. Sakai A (1993) Strategies for survival of plant cultured cells and meristems cooled
to 196 1C. In Cryopreservation of Plant Genetic Resources, pp. 1–16. Japan
Way (RBG Kew, United Kingdom); Dr. W. Holt (Institute of International Cooperation Agency.
Zoology, London, United Kingdom); Dr. S. Miyazaki and Mr. Steiner AM and Ruckenbauer P (1995) Germination of 110-year-old cereal and
A. Yamamoto (NIAR, Japan); Dr. B. Panis (Leuven, Belgium); weed seeds, the Vienna Sample of 1877. Verfication of effective ultra-dry storage
Dr. V. C. Pence (CREW, Cincinatti, United States); Dr. D. at ambient temperature. Seed Science Research 5: 195–199.
Sugawara H, Ma J, and Miyazaki S (eds.) (1999) World Directory of Collections of
Smith (CABI Bioscience, United Kingdom); Dr. S-H Tan (CBS,
Cultures of Microorganisms, 5th edition Japan: WFCC World Data Center on
Netherlands); Dr. L. Towill and Dr. C. Walters (NSSL, United Microorganisms.
States); Dr. M. Wood (London, United Kingdom); and Dr. X-Y. Touchell DH and Dixon KW (1994) Cryopreservation for seedbanking of Australian
Yang (XTBG, Peoples Republic of China). species. Annals of Botany 74: 541–546.
Towill LE and Forsline PL (1999) Cryopreservation of sour cherry (Prunus cerasus
L.) using a dormant vegetative bud method. CryoLetters 20: 215–222.
Vertucci CW and Roos EE (1990) Theoretical basis of protocols for seed storage.
See also: Biodiversity in Plant Breeding. Breeding of Animals. Plant Physiology 94: 1019–1023.
Captive Breeding and Reintroduction. Genetic Diversity. In Situ,
Ex Situ Conservation. Indigenous Strategies Used to Domesticate
Plants in Brazilian Amazon. Zoos and Zoological Parks