ON THE INHERITANCE OF COLOR IN A FRES~-WATERFISH.

APLOCHEIL17S LA TIPES TEMMICK AND SCHLEGEL. WITH

SPECI-4.L REFERENCE TO SEX-LINKED INHERITANCE
TATUO AIDA
.
Higher Technical School. Ky8ta Japan

Received May 27. 1921

TABLE OF CONTENTS
I.T.ODtrC.0. ................................................................... 5%
Materials and methods ........................................................... 555
Experimental results ............................................................. 555
Crosses between brown and red ................................................ 557
Crosses between brown and variegated red ...................................... 557
Crosses between variegated red and red ............................... 557
Crosses between white females and red males. ................................... 558
Crosses between white females and variegated red males .......................... 559
Crosses between white females and brown males ................................. 559
Crosses between blue and red ................................................. 561
Mating white with white ...................................................... 561
Crosses between blue females and white mafes ............................. 561
Crosses between blue and blue ............... ............................. 562
Crosses between brown and blue ............................................... 562
Crosses between brown and white .... ..................... 562
Crosses between red and white ................................................. 563
variegated red females and white males .......................... 564
............................................................. 564
Color-producing genes. ....................................................... 564
Triple allelomorphs........................................................... 565
Sex-linked inheritance ........................................................ 565
Location of the dominant gene, R,in the Y chromosome.......................... 569
Crossing over between the sex chromosomes ..................................... 570
SUMMARY ....................................................................... 572
CITED ..............................................................
LITERATURE 572
.................................................................... 573
POSTSCRIPT

INTRODUCTION

Aplocheilus lutipes (Japanese name ‘Medaka’) is a small fish of the

family Poeciliida: very common in our streams and paddy fields. Its
colored varieties are well known as ornamental fishes. Since. on accaunt
of their small size. a great many individuals may be bred within a small
space. they are very suitable objects for breeding experiments . Since
GENETICS 6 : 554 N 1921
 SEX-LINKED INHERITANCE I N A FRESH-WATER FISH 555

1913, experiments have been carried on to investigate the genetic behavior

of their body color, and some interesting results have been obtained so that
I think it will be worth while to report them here as a contribution to knowl-
edge about the genetics of fishes which has till now been very rarely
investigated .
MATERIALS AND METHODS

The brown-black variety was obtained wild from nature, while all others
were bought from a fancier. Among them, I was able to distinguish four
varieties, viz., orange-red; white, orange-red variegated with black, and
white variegated with black (plate 1, figures 1, 3, 5, 2).
I n nature this fish seems to live two years. The young, hatched out
in summer, spawn at the same season of the next year and soon die, so
that in autumn all fishes in nature are young and small. I n culture their
life may be prolonged one year more.
When the fishes are well nourished, they spawn every day early in the
morning, from the beginning of May to the end of August. According to
my observations, the number of eggs deposited daily averages 25; the
maximum was 71. The female carries the spawned egg-mass about half a
day, when not disturbed.
For the purpose of rearing the young, I stripped off the egg mass from
the female with the fingers, and then put the egg masses in a porcelain
vessel about one foot in diameter, one vessel containing not more than
five egg masses. The young hatch in about one week after spawning in
the hottest part of our summer.
As food for young just hatched, either powder of heated wheat grains
or any dried flesh is good, while fish which are somewhat grown, if nour-
ished with small earthworms (Tubificide), grow quite rapidly. The body
of a full-grown fish measures on the average 30 mm in length, without
the tail-fin. An aquarium of four square feet in surface area and measur-
ing one foot and a half in depth may contain 200 adult fishes in a healthy
condition, if proper care is taken to change the water.

EXPERIMENTAL RESULTS

For the sake of brevity, varieties which are more accurately described
as brown-black, blue-black, orange-red, orange-red variegated with black,
and white variegated with black, t d l hereafter be designated simply as
brown, blue, red, red variegated, and white variegated, respectively. The
number within parentheses denotes the year when the mating was made.
GENETICS6 : N 1921
 I PLATE
E ~ ~ A T OF O ~
Originally drawn to twice natural size, but reduced to one and three-fourths times natural
size in reproduction.
Figure 1.-Orange-red.
Figure 2.--White variegated with black.
Figure S.-White.
Figure 4.-Brown-black, wild form.
Figure 5.4range-red variegated with black
Figure 6.-BIue-black.

GENETICS6 : 556 N 1921

 SEX-LINKED
AIDA,TATUO, INHERITANCEI N A FRESH-WATER Fisn PLATE1

7
"g.1 nq. 2 Ry.3

R9.4
T
Rg. 5
T
(:ENETICS 6: N 1921
 SEX-LINKED INHERITANCE IN A FRESH-WATER FISH 557

Crosses between brown and red

Experiment 1
Brown males were mated to red females (1913). The F1progeny were
brown, and indistinguishable from the wild form. The result of their
inbreeding (1914) was as follows:
Brown Rea
Observed.. ............................................. 865 292
Expectation.. ............................................ 867.75 289.25

Experiment 2
Back-cross of experiment 1. Heterozygous brown females were mated
to pure red males (1914). The result was
Bvovm Red
Observed.. ............................................. 937 924
Expectation. ............................................. 930.5 930.5

Crosses between brown alzd variegated red

Variegated red individuals bought from a fancier were all heterozygous
for variegation, and produced red individuals when bred inter se. By se-
lective mating, I obtained in 1915 homozygous variegated red individuals
with which to perform further experiments.
Experiment 3
Homozygous variegated red males were mated to pure brown females
(1915). The offspring were indistinguishable from individuals of the
pure-bred brown strain. Their inbreeding (1916) gave the following result:
Brown Variegated red
Observed.. .............................................. 229 77
Expectation. ............................................. 229.5 76.5

Experiment 4
Back-cross of experiment 3. Heterozygous brown males were bred to
homozygous variegated red females (1916). The result was
Brown Variegated red
Observed. .............................................. 148 148
Expectation. ............................................. 148 148

Crosses betweem variegated red and red

Experiment 5
Homozygous variegated red females were mated to homozygous red
males (1915). The F1 offspring were all variegated red similar to the
variegated parents. The F2 offspring (1916) were as follows:
GENEncs 6 : N 1921
558 TATUO AIDA

Variegated red Red

Observed.. ............................................. 457 161
Expectation. ............................................. 463.5 154.5

Experiment 6
Back-cross of experiment 5. Heterozygous variegated red males were
bred to homozygous red females (1916). The result was
Variegated red Red
Observed ............................................... 216 200
Expectation. ............................................. 208 208

Though crosses reciprocal to the above six breeding experiments were

carried out, their details are omitted here because they have given exactly
similar results.
Crosses between white females and red males
White color is a sex-linked character, as explained below, and so the
sex of the progeny was carefully examined in all the following experiments.
To avoid error the examination as to sex was made after the fishes were
fully grown, females being distinguished as such by spawning and males
by having produced the long dorsal and anal fins.
Experiment 7
White females were mated to homozygous red males (1914). The F1
offspring were all red. The Fz offspring (1915) were as follows:
Red While
Observed.. ............................................. 217 76
Expectation. ............................................. 219.75 73.25

The result of sex examination was

Red White
& __L_
0 c? 0 #
Observed.. ............................... 41 76 43 0
Expectation ................................ 39 78 43 0

Experiment 8
Back-cross of the above experiment. Heterozygous red males were
bred to white females (1915). The result was
Red White
Observed.. ................................... 519 502
Expectation. .................................. 510.5 510.5

The result of sex examination was

Observed ..................................
-
0
2
Red
d
251
c_h_
0
197
While
d
1
Expectation. ............................... 0 253 198 0
 SEX-LINKED INHERITANCE I N A FRESH-WATER FISH 559

Crosses between white females and variegated red males

Experiment 9
Pure variegated red males were bred to white females (1917). The
F1 offspring were all variegated red. The Fzoffspring (1918) were
Variegated 7ed Variegated while Red White
Observed. ...................... 279 95 96 30
Expectation. .................... 281.25 93.75 93.75 31.25

The result of sex examination was

Observed.. ........ 46
-
0
Variegated red
r

90
d P
59
-
Variegated white
8
0
.
17
Red
-__.
3
37
3
-
P
17
White
d
0
Expectation. ....... 45.3 90.6 59 0 18 36 17 0

Crosses between white females and brown males

Experiment 10 A
Homozygous brown males were mated to white females (1915). The
F1 offspring were all brown. The Fz offspring (1916) were
Brown Blue Red Whife
Observed.. .................................. 248 57 53 21
Expectation. ................................. 213.12 71.04 71.04 23.68

The result of sex examination was

r
0
Bmm
$
. -0 Blue
- -8 9
Red
$ 0
White
13
Observed. ............. 77 147 56 0 9 37 19 0
Expectation. ........... 74.7 149.4 56 0 15.3 30.6 19 0

Experiment 11 A
Back-cross of experiment 10 A. Heterozygous brown males were mated
to white females (1916). The offspring were
Brown Blue Red White
Observed ........................ 387 317 253 246
Expectation ...................... 300.75 300.75 300.75 300.75

The result of sex examination was

Observed ..........
- 0
O
Brown
d
176
__?
0
214
Blue
8
1
__?
0
0
Red
8
135
__?
P
146
White
d
0
Expectation ........ 0 176 215 0 0 135 146 0

GENETICS 6 : N 1921
560 TATUO AIDA

In the last two experiments the deviation of the actual numbers from
expectation is rather large, so that I have repeated both experiments.
I n order to reduce as much as possible the influence of differential mor-
tality among the young of various colors, records were taken a t as early a
stage as was possible. First, at a very young stage, i.e., two or three days
after hatching, black and colorless young were distinguishable from each
other and were separated. Secondly, when the young fish had grown to
about 1 cm in length and the development of xanthophore had become
conspicuous, those previously classed as “ black’’ were distinguishable either
as brown or as blue and were separated accordingly, while those previously
classed as “colorless’’ were distinguishable as red or as white, and were so
classified.
Experiment 10 B
Repetition (1917-1918) of experiment 10 A. The first inspection of the
F2 generation gave
Black Colorless
O b s e ~ e ...............................................
d 2016 667
Expectation. ............................................. 2012.25 670.75

The black grew into

Browm Blue
Observed. .............................................. 779 244
Expectation. ............................................. 767.25 255.75

The colorless grew into

Red While
Observed.. ............................................. 329 113
. Expectation. ............................................. 331.5 110.5

The result of sex examination was

P
Brown
8
. -0 -Blue-8 0
Red
8 0
White
c?
Observed ............ 71 164 59 0 48 131 62 0
Expectation .......... 78.3 156.6 59 0 59.7 119.4 62 0

Experiment 11 B
Repetition (1918) of experiment 11 A. The first inspection gave
Black Colorless
Observed.. ............................................. 823 776
Expectation.. ............................................ 799.5 799.5

The black grew into

Brown Blue
Observed.. ............................................. 228 230
Expectation. ............................................. 229 229
 SEX-LINKED INHERITANCE IN A FRESH-WATER FISH 561

The colorless grew into

Red White
Observed.. ............................................. 237 222
Expectation.. ............................................ 229.5 229.5

Observed.. .......
-
The result of sex examination was

0
2
Brown

120
8
ch?
Q
114
Blue

0
d
- -
0
0
Red
8
71
Q
62
White
d
0
Exfiectation ........ 0 122 114 0 0 71 62 0

Crosses between blue and red

Experiment 1 2
Blue offspring of experiment 10 A were bred to homozygous red males
(1917). The progeny were
Brown Red
Observed.. ............................................... 222 234
Expectation ................................................ 228 228

The result of sex examination was

Observed. ..............................
_h_?

0
62
Brown

41
d
-
0
69
Red

48
d

Expectation. ............................. 51.5 51.5 58.5 58.5

Mati9ag white with white

Experiment 13
The solitary male which was found among the white offspring in experi-
ment 8 is the progenitor of all white males met with in further experi-
ments. He was mated to a white female (1916); the offspring, 149 in
number, were all white; 108 of them have grown to maturity, 66 being
female and 42 male.

Crosses between blue females and white males

Experiment 14
The object of this experiment was not only to study the genetic consti-
tution of blue individuals, but also to obtain blue males. Blue females,
derived from experiment 10 A, were mated to white males (1917). The
result was
Blue White
Observed ................................................. 146 151
Expectation. ............................................... 149 149

GENETICS6 : N 1921
562 TATUO AIDA

The result of sex examination was

Observed.. ...........................................
_h_
0
33
Blue

48
8
-
Q
51
Wkik

49
d

Expectation., ......................................... 40.5 40.5 50 50

Crosses between blue and blue

Experiment 15
Blue descendants of experiment 14 were bred inter se (1918). The
progeny were
Blue White
Observed ................................................ 618 200
Expectation............................................... 613.5 204.5

Crosses between brown and blue

Experiment 16
Pure blue females were mated to brown males (1919). The F1 offspring
were all brown. I n the Fz generation (1920), there were
B70W Rlue
Observed.. ................................................ 174 59
Expectation.. ............................................... 175 58

Crosses between brown and white

Experiment 17
White males were bred to pure brown females (1917). The FLoffspring
were all brown. The Fz offspring (1918) were a t an early stage
Black Colorless
Observed. ............................................. 1481 532
Expectation. ............................................ 1509.75 503.25

The black grew into

Brom Blue
Observed. ................................................. 477 150
Expectation.. ............................................... 470 157

The colorless grew into

Red Wk'hik
Observed.. ............................................. 208 69
Expectation. ............................................. 207.75 69.25

--
The result of sex examination was
B70W Blue Red Wkik
_rr__ cIc--r
0 c
9 Q d Q d O d
Observed.. ......... 173 88 0 79 78 34 0 31
Expectation .......... 174 87 0 79 74.6 37.3 0 31
 SEX-LINKED INHERITANCE I N A FRESH-WATER FISH 563

Experiment 18
Heterozygous brown males which were the F1 offspring derived from
experiment 17, were bred to white females (1918). The offspring were
a t an early stage
Black Colorless
Observed.. .............................................. 917 988
Expectation.. ............................................. 952.5 952.5

The black grew into

Brown Blwe
Observed.. .............................................. 354 339
Expectation. ............................................... 346.5 346.5

The colorless grew into

Red While
Observed ................................................ 363 378
Expectation. .............................................. 370.5 370.5

The result of sex examination was

Observed.. ............ 157

r___
0
Brown
d
0
--
0
0
Blae

148
8 0
165
Red
c?
0
_h_

0
0
White
8
126
Expectation.. ........... 157 0 0 148 165 0 0 126

Crosses between red and white

Experiment 19
White males were bred to homozygous red females (1917). The F1 off-
spring were all red. I n Fz (1918) there were
Red White
Observed.. .............................................. 766 264
Expectation.. ............................................. 772.5 257.5

The result of sex examination was

Red While
__I _h_
P d 0 8
Observed.. ..................................... 87 42 0 33
Expectation. ..................................... 86 43 0 33

Experiment 20
Heterozygous red males produced as F, ofispring in experiment 19 were
bred to white females (1918). The offspring were
Red White
Observed.. ................................................ 330 324
Expectation. ................................................ 327 327

The result of sex examination was

Observed.. ......................................
-
9
135
Red
3
0
-
9
0
White
6 ”
153
Expectation.. ..................................... 135 0 0 153

GENETICS6 : N 1921
564 TATUO AIDA

Crosses between variegated red females and white males

Experiment 21
White males were mated to homozygous red variegated females (1917).
The Fl offspring were all red variegated. I n the Fz generation (1918),
there were
Variegaled red Variegated whik Red Whtte
Observed.. .................... 157 50 49 18
Expectation.. ................... 154.125 51.375 51.375 17.125

The result of sex examination was

Observed.. ...........
Variegakd red
c _ _
0
76
8
35
-- -
Variegated while
0
0
8
34
0
27
Red
3
5
0
0
White
3
6
...........
Expectation. 74 37 0 34 21.4 10.7 0 6

DISCUSSION

Color-producing genes
White is recessive to any other color and breeds true (experiment 13).
Red and-white, when crossed, segregate out, each to its respective original
color in the Fz generation, according to the simplest Mendelian ratio (ex-
periments 7, 8, 19, 20), from which we see that red color is produced by a
single dominant gene which I will call R. The cross between brown and
red gives also in Fz the original colors according to the 3 :1 ratio (experiment
1)) from which we see clearly that brown color has one more gene, B, in
addition to the red-producing R. That the genetical constitution of brown
should be BR is confirmed by the cross of brown with white, because in the
Fz generation four varieties, brown, blue, red, and white, are produced
according to the dihybrid ratio, 9 :3 :3 :1 (experiments 10, 17). The same
fact is more directly proven by the result of experiment 12 where the cross
blue X red gave rise to brown offspring. Blue individuals, which originated
in my cultures, possess the gene B, which is also contained in brown ones,
as shown by the results of experiments 14, 15 and 16. TOYAMA (1916)
and ISHIWARA (1917) have reported results exactly similar to mine derived
from crosses between brown, red, and white.
Variegation is produced by another gene, B'; this is clearly proven by
the cross between variegated red and red (experiments 5, 6), where we see
the monohybrid segregation, as well as by the cross between variegated red
and white (experiments 9, 21) where we see the dihybrid segregation con-
cerning the genes B' and R.
 SEX-LINKED INHERITANCE I N A FRESH-WATER FISH 565

The action of the above-mentioned color-producing genes may be re-

garded as follows:
B is a gene able to produce black pigment or melanophore uniformly
throughout the whole body;
R is a gene able to produce yellow pigment or xanthophore;
B' is a gene able @o produce black pigment only partially in the body.
I n accordance with the foregoing hypotheses, the genetic constitution of
the several color varieties, when homozygous, would be: brown, BBRR;
blue, BBrr; red, bbRR; variegated red, B'B'RR; variegated white, B'B'rr;
and white, bbrr.
Triple allelomorphs
The three colors, brown, variegated red, and red, form a triple allelomor-
phic series. Any two of them, when crossed, show in F2the 3 :1 segrega-
tion (experiments 1, 3, 5). Brown is dominant to variegated red as well
as to red, while variegated red is dominant to red. When brown and red
are crossed, the Fz off spring produce brown and red in the ratio 3 :1, and
do not contain any variegated red, which is the characteristic behavior
of multiple allelomorphs.
A similar mode of inheritance was described by IBSEN (1919) in respect
to the black, tortoise, and red colors in guinea-pigs, which are produced,
respectively, similarly as in our three varieties, by complete extension,
partial extension, and non-extension of black pigment.
Sex-linked inheritance
The results of experiments 7 to 11 as well as 17 to 21 attract our atten-
tion on account of a remarkable manifestation of sex-linked inheritance.
When a white female (rr) is mated to a male possessing the R gene in a
homozygous state, all Fz white offspring. (rr) as a rule are female. Thus,
in experiment 8 all white individuals except one, were female, and in the
cross, white female X variegated red male (experiment 9)) all white and
variegated white individuals were female, and in the cross white female
X brown male (experiments 10, 11)) all blue and all white individuals,
except one of each, were fema1e.l
On the contrary, when a white male is mated to a female possessing the
R gene in a homozygous state, all Fz white, blue or variegated white indi-
viduals as a rule are male (experiments 17, 19, 21).
All these peculiar results may be explained in the following way: First,
the sex inheritance of the fish under discussion belongs to the Drosoph-
'That all whites are female without exception has been noticed by TOYAMA
(1916), but
I S ~ W A (1917)
R A had both male and female whites in his experiments.

GENETICS6 : N 1921
566 TATUO AIDA

ila type, XX-XY, i.e., to that type in which the female is homozygous
and the male heterozygous for the sex (or X) chromosome. HUXLEY
(1920, page 270) recently assumed the same type of sex inheritance to
occur in the fish, Giardinus. Second, the red-producing gene R is located
in the chromosomes X and Y .
The male homozygous for red color has one R gene contained in t h e X
and another contained in the Y chromosome, which I represent by R,
and R,, respectively; while the female homozygous for red color has one
R gene contained in each of the two X chromosomes which I represent
by Rx.
As a critical test for the above view the results of experiment 8 may be
cited. I n this experiment the male FI offspring derived from the cross
white female X homozygous red male were mated to white females. Had
the sex been inherited according to the Abraxas type, in which the female
is heterozygous and the male homozygous in respect to the sex-determin-
ing genes, we should expect to have as the result of the experiment, red
females and males, and white females and males in equal numbers. The
actual result was quite different; all whites, 197 in number, were female
except only one individual, while all reds, except 2 out of 253, were male.
When we assume the Drosophila type in our case, these results are in per-
fect agreement with the theoretical expectation, as we shall see in the follow-
ing diagram.
rxrx (white 0 ) X RxR, (red$)
I
I I
Rxrx (red 0 ) rxRy ( r e d g ) X rXrx (white 0 )
I
I I
rxr, (white 0 ) rxRy (redd')
(all white) (all red)
As above stated, the mode of sex inheritance in our fish belongs to the
Drosophila type. The Y chromosome in our case differs however from
what we see in Drosophila, inasmuch as it contains the dominant gene R.
The production of red (dominant) males from the cross between white
(recessive) female and red (dominant) male, as seen in the above diagram,
would be otherwise quite unexplainable.
All other experimental results concerning sex linkage are equally well
explainable according to the above hypothesis. Below I will give in the
form of diagrams the theoretical expectation and the results actually ob-
tained in experiments 7, 9, 10, 11, 17, 18, 19, 20, and 21.
 SEX-LINKED INHERITANCE I N A FRESH-WATER FISH 567

Experiment 7
RxRy (redd') X rxrx (white 9 1
1

I I
Rxrx (red 9 1 X rxRy (redd')
I
I I I I
Rxrx (red 9 rxr, (white 9 ) RxR, (redd') rxRy (redd')
Red Whdk
_h_ ch_
O d 0 3
Theoretical sex ratio.. .............................. 1 2 1 0
Observed.. ....................................... 41 76 43 0

Experiment 9
B'B'RxR, (variegated red 8) X bbrxr, (white 9 )
I
I I
B'bRXrx(variegated red 9 ) X B'brxRy (variegated r e d s )
I
I I
B'B'R,R,

2B'bR,rx
'I
B'B'rXRy (variegated
2B'bRxRy red C? )
2B'brxR,
B'B'R,rx (variegated
]
red 9 )
- -
Variegated red Variegated while
bbR,r, (red 9 ) bbrxvx (white 9 )

bbR,R, }(redc?)
bbrxRy

__L_
Red
cc_
Whik
0 3 0 3 0 3 0 3
Theoretical sex ratio. ... 1
' 2 1 . 0 1 2 1 0
Observed.. ........... 46 90 59 0 17 37 17 0

Experiment 10
BBRxR, (brownd') X bbrxYx (white 9 )
1
I I
BbR,r, (brown 9 ) X BbrxRy (brownc?)
I
I I I I
BBRxRy
BBrxRy
2BbRxRy
2BbrxRy

2BbRxrx
I
(brownc? )
2 Bbr, r, 9)
BBrxYx](bhe
bbR, Y, (red 9 )

bbR,Ry
bbr,R,
bbr,r, (white 9 )

GENETICS6 : N 1921
568

--
TATUO MDA

Theoretical sex ratio. ....1

P
Brown
d
2 1 0
O
Elare
d
ch_
O
1
Red

2
d
-Q
1
White
d
0
Observed*. ............. 148 211
11.5 0 57 168 81 0
*Sum of the results of two experiments, 10 A and 10 B.

Experiment 11
B6QY (bromc?) X bbr,r, (u.hiteO)
f
I I I 1
Bbr,R, (brown$) Bbrxrx (blueP) bbrXRs (red$) bbr,r, (white 1
Brown Bise Red White
Theoretical sex mtio.

ments, 21 A and 11 B)
. ..
Observed (the sum of the
results of two experi-
I

2
all 3

286 328
7d Q
all 0

1.
-
0
all 3

206
d Q
208
all 0

0
d *

Experiment 17
BBRx& (6rown9f X 6&rXry
( w ~ i ~ ~ ~ ~
I
i I
BbRxr, (brown9) X BbRxry (brown81
I

Theoretical sex ratio. .... 2

__c_
P 5-
1
- c

0
O c i r
1
ch_
Q
2 1
d
0
-
O
1
d

Observed.. ............ 173 88 0 79 78 34 0 31

Experiment 18
BbRxry (brown$) X bbr,~, (white9)
I
i f -i-- f
BbR,r, (brown 0 1 S&gY(blue$) bbRxrx (red 9 1 bbrxry (white#)

Theoretical sex ratio. ....

Observed.. ............157
- --
0
EFm
all 0
3
0 *
?
0
Bke
a w
d
148
O
165
Ked
afl0
d
0
?
0
Whike
allo*
___c_
d
126
 SEX-LINKED INHERITANCE IN A FRESH-WATER FISH 569

Experiment 19
RxRx (red91 X rxry (white$)
I
I t
Rxrx (red 9 ) X Rxry (red$)
I
1 I I I
RxRx (red 0 )

Theoretical sex ratio.

R,r, (redo)

.........................
Rxry (redd')
-
O
2
Red
d
1
rxry (whitecP)
p
O
0
h?
Whib
d
1
Observed.. ................................. 87 42 0 33

Experiment 20
Rxry (red$) X rxrx (whiteQ)
I
I I
R,r, (redQ) r g , (white$)
Red Wh&3
Theoretical sex ratio. ......................... all 9 all8
_I___ p__
0 d 0 . 9
Observed. .................................. 135 0 0 153

Experiment 21
B'B'RxRx (variegated red P) X bbrxry (white$)
I
i t
B'bRxrx (variegated red 0 ) X BfbRxry(Variegated red&
II
I I
(variegated bbR,R, bbrxry(whited')
white$) bbRxrx](red 0 )

2BfbRxrx bbRxry (red$)

B'B'Rxry (variegated
}
2B'bRxry red$)

Theoretical sex ratio. .....

Variegafedred
r___
O
2
d
1
-
Variegated White
0
0
6
1
cIc_

2
0
Red
8
1
-
0
0
White

1
8

Observed ............... 76 35 0 34 25 5 0 6

Location of the dominant gene R in the Y chromosome

One of the notable features in the sex-linked inheritance above described
is the fact that the Y chromosome contains the dominant gene R. All
GENETICS6 : N 1921
5 70 TATUO AIDA

red males bought from the fancier were homozygous for that color. That
the wild brown male contains the R gene in its Y chromosome is to be seen
clearly from the results of experiments 10 and 11. Since the Y chromosome
is inherited by the male, exclusively, and consequently goes from the father
t o his son only, all males contained in the progeny of any mating to which
either a red or a brown male is a party, possess the gene R, i.e., they are
red, red variegated, or brown; whilst all individuals lacking the gene R,
Le., white, white variegated, or blue, are female (experiments 7, 8, 9, 10
and 11).
According to the investigations hitherto reported, the Y chromosome
in the Drosophila type and also the W chromosome in the Abraxas type
(corresponding to the Y chromosome in Drosophila) are known to possess
no dominant gene, so that all characters contained therein are recessive
in genetic behavior. The fact that in our fish this chromosome carries
a dominant gene instead of a recessive is very interesting, and will bring
out a certain important significance of its existence.

Crossing over between the sex chromosomes

No example of crossing over between a sex chromosome and any other
chromosome has been reported till now, and it has been supposed that, be-
cause this is so in the heterozygous sex, the X and Y chromosomes are not
exactly homologous to each other so as to permit the crossing over of genes
between them. I n my experiments the crossing over of the R gene be-
tween them seems not to be very rare. I n experiment 8, in which all red
were expected to be male and all white to be female, two reds out of 253
were female (Rxrx) and one white out of 198 was male (rxry). I n
experiment 11 A, where all blues were expected to be female, one out of
215 was male (Bbrxry); and in experiment 11 B, where all browns were
expected to be male, two out of 122 were female (BbR,r,).
The production of such unexpected individuals is, in my opinion, best
explained by assuming the crossing over of the R gene from a U to an X
chromosome. One might perhaps think that it may be caused by the
process of non-disjunction discovered by BRIDGES(1916). But if this
were the correct explanation, all exceptional females would have three
sex chromosomes, viz., two X and one Y chromosome, and all exceptional
males would have only one X chromosome. Thus the constitution of the
red female in experiment 8 should be r,r,R,, that of the brown female
in experiment 11 B should be Bbr,r,R,, that of the white male in ex-
periment 8 should be r,O, and finally that of the blue male in experi-
ment 11 A should be Bbr,O. The results of experiments 17, 18, 19,
 SEX-LINKED INHERITANCE I N A FRESH-WATER FISH 571

20, and 21, in which the male offspring of the exceptional white male de-
rived from experiment 8 were mated to various females, would then have
to be explained by assuming the constitution of the original male to have
been r,O.
In order to settle the question, which of the two views,-crossing over
or non-disjunction,-will better explain the production of unexpected in-
dividuals, the result may be cited of a mating between heterozygous red
females and males (experiment 22, not previously described in this paper).
The red mothers in this experiment were F2 offspring from the cross
white 9 X brown 3 (experiment 10 B) and should have the constitution
R, r, , while the red fathers were F2 offspring from the cross brown 9
X white 8(experiment 17) and should have the constitution either R,r,
or R,O, according as we’adopt the view of crossing over or that of non-
disjunction. The result of the cross was

Experiment 22 (1919)
Red 9 (Rxrx) X R e d 3 (RXryor R,O)
I
i I
Red White
Observed ...................... 451 149
Expectation ................... 450 150

The result of sex examination in 1920 was

Observed .....................................
c____
0
146
Red
d
57
-
O
2
White
d
80
Expectation. ................................... 135.4 67.7 0 82

Here also two unexpected white females were produced. If we consider

the constitution of the red male to be R,O, the production of the white
females is not explainable, because the white female should have two
X chromosomes, i.e., should contain rxrx, but in the cross of R,r, fe-
male with RO , male there would be but one single gene r. By assuming
the constitution of the red male to be Rgry and supposing moreover that
the crossing over takes place between the chromosomes R, and r,, so
that chromosomes r, and R, are thereby formed, the production of
r,r, may clearly be explained. We are led consequently to the conclusion
that the production of the exceptional red females and the white male in
experiment 8 is due, not to the process of non-disjunction, but to the cross-
ing over of the R gene between the X and the Y chromosomes. As to
exceptional individuals produced in experiment 11 my endeavors to
GENETICS 6 : N 1921
572 TATUO AIDA

determine their origin have been thus far in vain, though their production
by crossing over seems to me to be highly probable. At present, my data
concerning this question are yet very scanty, and I am unable to make
any further inference in this respect, but I hope to be able to give much
more detail in a future paper, because the experiments for the purpose are
now in progress.
SUMMARY

1. I n Aplocheilus latipes Temmick and Schlegel, the wild form is brownish

black but four novel color varieties are known, viz., orange-red, orange-
red variegated with black, white, and white variegated with black.
2. The genetic consequences of a mating between any of the three forms,
brown, red, and white, are quite simple. Brown is produced by the genes
B and R, red by the gene R and white contains no dominant gene.
3. I n the Fz generation of the cross between brown and white a new vari-
ety of a blue-black color first made its appearance. It is characterized by
the dominant gene B.
4. Variegation is produced by the gene B'. The three genes, B, B',
and the recessive b, form a series of triple allelomorphs.
5 . Sex inheritance is of the Drosophila type, XX-XU, i.e., the female
is homozygous and the male heterozygous in respect to the sex-deter-
mining genes.
6. I n the male homozygous for the R gene, one R is carried by the Y
chromosome. Since this chromosome is transmitted from male to male,
whatever the genetical constitution of the female may be, all male progeny
contain the R gene, and consequently neither white nor blue males are nor-
mally produced.
7. Crossing over of the R gene between the X andtheYchromosomeswas
found not rarely in the offspring of back-crosses between heterozygous
brown or red males and white females and also matings of heterozygous red
males with heterozygous red females.
LITERATURE CITED
C. B., 1916 Non-disjunction as proof of the chromosometheory of heredity. Genetics
BRIDGES,
1: 1-52, 107-163.
HUXLEY,J. S., 1920 Note on an alternating preponderance of males and females in fish, and
its possible significance. Jour. Genetics 10: 265-276.
IBSEN,
H. L., 1919 Tricolor inheritance. 4. The triple allelomorphic series in guinea-pigs.
Genetics 4: 597-606.
ISEIIWARA,
M., 1917 On the inheritance of body-color in Oryzias latipes. Mitteilungen aus der
medizinischen Fakultat Ryushu 4: 43-51.
TOYAMA,
K., 1916 Itiniino Mendel-Seisitu ni tuite. [On some Mendelian characters.] Nippon
Ikusyugakkukwai H6koku [Report of Japanese Breeding Society] 1: 1-9.
 SEX-LINKED ~ ~ E ~ T A NI NC AE FRESH-WATER FISH 573

PoSTSCRIPT.-The manuscript of the present paper was sent to Dr.

SHULLa t the end of April 1921. On the last day of May, “Science,” Vol.
53, No. 1371, published April 8, came to my hands, and on reading there
the paper of Dr. CASTLE,entitled “A new type of inheritance” (pp. 339-
342), I first became acquainted with the investigation of Dr. SCHMIX)T of
Copenhagen, concerning the “Millions fish,” Lebistes retkzdatzcs. Though
I am not yet able to get his original paper, published in “Comptes rendus
des Travaux du Laboratoire de Carlsberg,” Vol. 14, No. 8, 1920, his ob-
servations as described in “Science” are as follows: A conspicuous black
spot occurring on the dorsal fin of the male in one race of this species is
transmitted exclusively from father to son, regardless of the mother’s
ancestry. Several cross-experiments have led SCHMIDT to the conclusion
that the dominant gene causing that sex-linked character must be located
in the Y chromosome, which has been considered till now to be quite
“empty,” especially in Drosophila.
The discovery of Dr. SCHXIDTconsequently accords well with what I
have observed in ~ ~ ~ c ~Z ~u ~L~ ~zand
e cs , shave described in the present
paper, though the two cases differ in several minor details.2 It is very
interesting that the new mode of inheritance of exactly similar type was
found by Dr. SC-T and me quite independently from each other at
almost the same time, especially so because both relate to the same animal
group-the fishes,
~ED~TORUI. NOTE. It should be pointed out, in justice to the author, that his observations
go beyond those of SCHMIDTin the important respect of showing the occurrence of crossing over
between the X and the Y chromosomes.-W. E. CASTLE.

GENETICS
6 : N 1921