genes that affect early developmental stages of drosophila melanogaster

(W. G. K~RC~OFF Laboratories, California Institute of Technology, Pasadena, California.)

GENES THAT AFFECT EARLY DEVELOPMENTAL STAGES OF DROSOPHILA MELANOGASTER.

By

T~. DOBZHA~SKY and F. N. DUNCA_W.

With 7 figures in the text.

(Eingegangen am 18. April 1933.)

Table of contents. p a g e

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109 Technique . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111 Weight of chubby and wild-type . . . . . . . . . . . . . . . . . . . . 113 Time of appearance of the effect of chubby . . . . . . . . . . . . . . . . 114 Rate of growth of chubby and wild-type larvae . . . . . . . . . . . . . . 116 Absence of a maternal effect . . . . . . . . . . . . . . . . . . . . . 117 On the nature of the chubby character . . . . . . . . . . . . . . . . 119 Growth in rudimentary-12 larvae . . . . . . . . . . . . . . . . . . 120 Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 127 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 Zusammenfassung . . . . . . . . . . . . . . . . . . . . . . . . . . 129 Literature cited . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129

Introduction. The known number of muta t ions affecting the s tructure of the adul t

Drosophila melanogaster runs into hundreds. I n a sense the same is t rue for the earlier developmental stages (eggs, larvae and pupae). Lethals, which seem to be the most frequent kind of mutat ions, cause dea th of the organism at stages preceding the adult . The nature of the changes produced by lethals is, however, mos t ly unknown, and even the stage at which the death takes place is determined for a relat ively small number of lethals (LI 1927, SrV~TZEv-DoBz~A~SKY 1927). The morphology of the individuals dying in early stages of development (especially in the egg stage) consti tutes a mos t promising field of investigation. A t present this field remains a terra incognita.

The number of known factors producing visible changes in the structure of eggs, larvae and pupae is ve ry small. This is pa r t ly due to the simplicity of the external morphology of these stages as compared with the adul t stage, since in general the chances of finding a clear

110 Th. Dobzhansky and F. N. Duncan:

morphological difference between any two forms are proportional to the number of complex and diversified structures possessed by the object in question. The major reason is, however, tha t the at tention of the Drosophila workers up to the present has been devoted almost exclusively to a study of the adult flies.

The appearance of l>rosophila eggs is influenced by several factors. The eggs of females homozygous for singed are shorter than the wild-type eggs and have different shaped filaments (Ylom~ 1922). Eggs homozygous for Star develop a black spot in the region of mieropyle and die (SIwRTzEv- DOBZ~AWSXY 1927). The deficiency known as Minute-1 changes the shape of the eggs and the color of their yolk (LI 1927). Finally, Wx~RE~ (1924) described variations of the size and shape of the eggs caused by special modifying factors 1. Several genes are known which produce tumorlike malformations in larvae and pupae (BRIDGES 1916; STXRK 1918, 1919; ST~RK and BRII)GES 1926). Some of these genes act as lethals while others cause ,,benign" tumors. The length of the larval and pupal development is modified by many genetic factors (for example, the Minutes, SOHVLTZ 1929). ALPATOV (1930) found tha t the gene vestigial, which has a manifold effect on the adult, delays also the rate of growth of the larvae.

The recessive gene, known as chubby, discovered by Dr. C. B. BRIDQES (MoR*A~, B~rDGES, STURTEVAN~ 1925, p. 222), is located a t about 72 in the second chromosome (Mo~ xw 1932, p. 76). Mature larvae and pupae homozygous for chubby are shorter and broader than normal (Figs. 1 and 2). The adult chubby flies are also shorter and stockier, and the abdomen is more rounded than in wild-type, but the difference between adult chubby and wild-type flies is generally less striking than that between chubby and wild-type larvae and pupae. The classification of adult chubby is somewhat hazardous, while mature larvae and pupae can be recognized readily.

Another factor affecting the shape of larvae and pupae in a manner similar to chubby was found in the stock of rudimentary-12 flies by Mrs. N. P. SrWRTZEv-DoBZHA~SKY (rudimentary-12 is a sex-linked recessive producing shortening of wings and disarrangement of the hairs

1 Warren studied the eggs laid by various mutant races and often found them different. He then crossed the mutants in question to wild-type, and observed the segregation taking place in F 2 with respect to the characteristics of the eggs and of the adults. The result was that the mutants recovered in F~ laid eggs which were frequently somewhat different from those laid by the same mutants, before crossing. Warren concluded that the egg-shape is not determined by genes responsible for the production of the characteristics of the adult mutants, but rather by independent modifiyng factors. This conclnsion seems to us not entirely proven by his data. I t remains possible that the same mutant genes which affect the structure of the adult produce some effect on the egg-shape, but this effect is obscured by that of the independent modifiers (comp. DOBZ~SKY 1927).

Genes that affect early developmental s~ages of Drosophila melanogaster. 111

on the wing-margin). This factor is sex-linked, and is apl~rently localized rather close to rudimentary-12 (see below).

1 X Fig. 1. N e w b o r n l a r v a e of w i l d - t y p e (left) a n d F ig . 2. lYIature l a r v a e 0s w i l d - t y p e

of ch, u b b y ( r igh t ) . The scale to t h e le f t (left) a n d of c h u b b y ( r igh t ) . T h e scale r ep r e sen t s 0,5 of a m m , r ep re sen t s 2 m m .

The present contribution is devoted to the s tudy of the effects of chubby gene and of the factor associated with rudimentary-12. The authors wish to express their gratitude to Mrs. N. P. SIVERTZEV-DOBZHAN- SKY and Mr. J. BOmbER for their valuable ~ssistance in this work.

T e c h n i q u e . An investigation of the effect of any gene involves a comparison

of individuals carrying that gene with wild-type or with other mutants. Such a comparison can be accurate only in case the individuals studied differ from each other solely in the gene under consideration, and are isogeneous in respect to all other genetic factors. The stocks bred in laboratory do nob always comply with this requirement, since they frequently differ from each other in minor modifying factors which may affect any given character. Hence, before an accurate comparison can be made the stocks involved should undergo a procedure by which a desired degree of isogeneity is obtained.


Chubby flies were crossed to wild-type from the ,,inbred Oregon" strain. F~ was raised, and chubby flies which appeared there were selected using the characteristics of the adult as the criterion. The chubby flies were again crossed to wild-type from the original inbred strain, and this procedure was repeated five times. I t is higly probable t ha t the chubby strain obtained as the result differs from the ,,inbred Oregon" strain only in the chubby gone.

Vermilion rudimentary-12 males were crossed to ,,inbred Oregon" wild.type females. The F 1 females were back-crossed to wild-type males from the original strain. In F~ rudimentary-12 (not vermilion) males were selected and again crossed to wild-type females. This was done six times. A homozygous rudimentary-12 stock was then established. The larvae and pupae were short and stout. This suggests tha t the characteristics of the larvae and pupae are either due to a manifold effect of the gene rudimentary-12 itself, or to the effect of a separate locus closely linked with rudimentary-12. The lat ter possibility is made more probable by the fact tha t six other allelomorphs of rudimentary (rudimentary-7, rudimentary-14, and four more undescribed allelomorphs obtained in X-rays experiment by A. H. STV~TEVANT and T~. DOBZ~AW- SXY) were tested, and found to have no appreciable effect on the shape of the larvae and pupae either by themselves, or in crosses to rudimentary-12. All of these rudimentary allelomorphs have a stronger effect on the wing-size than has rudimentary-12.

The technique of the s tudy of the growth rate of Drosophila larvae has been very carefully worked out by ALPATOV (1929, 1930). We followed ALPATOV'S procedure with minor variations. Well fed flies were placed in culture bottles without food but with walls covered with moist filter paper. Paper spoons with ordinary Drosophila food were then inserted into the bottles. As soon as the flies laid a sufficient number of eggs on the surface of the food in the spoons, the latter were removed and fresh spoons substituted in their place. Spoons with eggs were kept in an incubator a t 270 C, and controlled every two hours under a binocular. The larvae hatched during each two hour period were placed in a separate PEr~I dish with food, and were then kept in the same incubator. The age of the larvae (after the emergence from the eggs) was, thus, known with an error not exceeding -t- 1 hour. At desired intervals some of the larvae were removed from the PET~I dishes, fixed, and preserved in 70 % alcohol. The fixation is performed by rapidly pouring boilling water on the larvae placed in a small amount of moisture in a watch glass. Most of the larvae die in a perfectly extended condition, a few of them are somewhat contracted, and these few were discarded.

Eggs, larvae and pupae were measured under a microscope in terms of the units of an ocular micrometer. For measuring eggs a relatively high magnification was used (one division of the ocular micrometer equals

Genes that affect early developmental stages of Drosophila melanogaster. 113

8,9 micra). For young larvae a lower magnification was used (one division 31,2 micra), and mature larvae and pupae were measured under a

still lower magnification (one division : 50,8 micra). The length and the maximum width of the eggs, larvae and pupae was measured.

In the following tables all the data are shown in millimeters. For each age the mean value with its mean error (M ~: m), the standard deviation (o), the coefficient of variation (C), the limits of variation (Lira), and the number of individuals measured (n) are given.

The instar of larvae was determined by observing the sizes of the month armature. As shown by ALrATOV (1929) the mouth armature does not grow between the moultings, and the dimensions of the mouth armature in the suecesive instars do not vary transgressively.

Weight of chubby and wild-type. Chubby larvae and pupae are shorter and broader than wild-type

(Figs. 1 and 2). The first question to be asked regarding the action of the chubby gene is wheter this gene affects only the shape or both shape and weight of the organism ? This question can be answered by weighing chubby and wild-type larvae and pupae of similar age.

Living eggs, larvae and pupae were extracted from the food, placed in watch glasses, and washed with water to remove particles of the food and of yeast adhering to the body surface. The excess of water was removed by gently rolling the objects on the surface of dry filter paper. Then the objects were transferred to a microbalance, and weighed 1 The results are shown in Table 1.

Table 1. Weight of chubby and wi ld- type (in rag). Chubby Wild-type

Object weighed

100 eggs 20 larvae 20 larvae 10 larvae 20 larvae 20 pupae

2Lge in hours

1;- 291/2 52 741/2

Total Weight of weight 1 specimen

1,49 0,0149 1,16 0,0580 2,69 0,1345 8,33 0,8330

29,89 1,4945 24,70 1,2350

Object weighed

100 eggs 20 larvae 20 larvae 10 larvae 20 larvae 20 pupae

Age in h o u r s

1;- 29 521/2 741/2

Total Weight of weight 1 specimen

1,50 0,0150 1,19 0,0595 2,48 0,1240 9,43 0,9430

31,87 1,5935 25,51 1,2755

The weights of the eggs and of the young larvae of chubby and wild- type are approximately alike. Mature chubby larvae and pupae are slightly lighter than wild-type, but the difference is very small. One m a y conclude tha t the gene chubby influences only the shape of the body, leaving the weight (and, presumably, the volume) unaffected, except in the later stages of development.

1 We are indebted to Dr. K. V. TH~AN~ for his kind help in performing these weighings.

W. Roux' ~rehiv f. Entwicklungsmeehanik. Bd. 130. 8


The t ime of development in chubby and wild-type is the same. In both pupation takes place between 80 and 100 hours after the emergence from the egg. The adult emerges from the pupa between 90 and 110 hours after pupation (at 270 C). The length of the embryonic development was not studied in detail, but it seems to be similar in both forms.

Time of appearance of the effect of chubby. I t is logical to inquire at what stage in development the effect of

chubby first becomes noticeable. The effect of a gene-difference on an organ m a y appear simultaneously with the appearance of the rudiment of the organ itself, or it may become apparent only later. In the first case one may conclude tha t the gene-difference 1 affects the formation of the organ, or the processes preceding thereto. In the second case we are dealing with a modification of the development of the organ already laid d~wn (P-,,Jn~TCH~XO 1929). Cases in which a gene-difference affects both the formation and the development of organs are, of course, also to be expected.

Eggs laid by homozygous chubby and by homozygous wild-type females were studied. The results are shown in Table 2. The length of the chubby and of the wild-type eggs is the same. The width seems to be greather in wild-type than in chubby (the reverse is true in larvae and pupae, see below), but the difference is not statistically significant.

Length { Chubby . . . . Wild-type . . .

Chubby . . . . Width 1 W i l d - t y p e . . .

Table 2. Size of the eggs

l~I•

0,556• 0,557-4-0,0016

0,209=l=0,0006 0,212~0,0009

0,025 0,022

0,009 0,013

(in mm).

c

4,5 4,0

4,1 6,1

Lima

0,481--0,614 0,498--0,623

0,187--0,240 0,187----0,267

200 200

200 200

The length of newborn chubby larvae is less than tha t of wild-type larvae (Tables 3 and 4, the difference is 0,052 ~: 0,011). The width of newborn chubby seems to be, to the contrary, higher than in wild-type (Tables 5 and 6), but the difference is not significant. At the age of four hours, however, the difference in width becomes fully significant.

x We speak here about the effect of "gene-differenees" rather than about the effect of the genes themselves. Comparing an organism carrying the gene A with an organism carrying the allelomorphic gene a one may obtain information regarding the effect of the gene-differenee A-a, but not regarding the effects of the genes A and a themselves. Much confusion would be avoided if all biological writers would realize this simple fact. The effect of a gene can be studied only by comparing organisms carrying this gene with organisms in which this gene is absent (deficiency). I t seems to us highly significant that, in Drosophila, individuals homozygous for any known deficiency die during the earliest stages of development (LI 1927).


One is jus t i f ied in concluding t h a t the difference be tween c h u b b y and wi ld - type becomes a p p a r e n t in the newborn larvae.

H

III

Table 3. L e n g t h of the b o d y of c h u b b y l a rvae .

Age in hours

Newborn 4 8

I 12 16 20 24 20 24 3O 36 42 48 42 48 54 6O 66 72 78 84 9O 96

M •

0,777:3=0,009 0,920:J::0,009 1,037:3=0,009 1,061 {=O,OO9 1,211 :j::O,O14 1,336:J::0,011 1,308 - - 1,426:J:0,026 1,546~0,021 1,822i0,016 2,026~0,028 2,232~0,026 2,169 - - 2,656:b0,037 2,966:j::0,062 3,227 :L0,064 3,624~0,052 3,692:J::0,029 3,850~0,040 3,946~0,039 3,734~:0,046 3,596:J:0,035 3,697-V0,046

a=d :

0,089 0,071 0,074 0,056 0,082 0,047

o,T 2 0,128 0,098 0,168 0,137

o,~7 0,371 0,376 0,312 0,174 0,240 0,238 0,278 0,207 0,276

c I

Lim

0,562--0,998 0,749--1,092 0,874--1,217 0,967--1,217 1,030--1,373 1,248--1,404 1,279--1,373 1,248--1,622 1,373--1,810 1,560--2,028 1,685--2,402 1,903--2,434 2,083--2,286 2,402--2,964 2,134--3,607 2,489--3,861 3,048---4,470 3,251--4,013 3,150---4,267 3,404--4,420 3,404--4,166 3,150--4,115 3,099--4,267

100 64 68 36 36 18 7

20 36 36 36 27

3 16 36 35 36 35 36 36 36 36 36

Table4. L e n g t h of the b o d y of w i l d - t y p e l a rvae .

II

III

Age in hours 1M: • m a = • C Lira n

Newborn 4 8

12 16 20 24 20 24 30 36 42 48 42 48 54 60 66 72 78 84 9O 96

0,829~0,006 0,975~=0,009 1,114=L0,009 1,192~0,007 1,304:t=0,013 1,468~0,008 1,457 - - 1,552=1=0,026 1,77010,024 2,229~0,019 2,560=i=0,013 2,646~0,038 2,286 - - 3,089+0,045 3,439+0,029 4,336~0,037 4,465i0,031 4,80410,035 4,530i0,050 4,953:J:0,039 4,823~0,047 4,583~0,049 4,705~0,058

0,065 0,073 0,075 0,042 0,080 0,045

0,~7 0,142 0,105 0,077 0,166

0,~1 0,179 0,213 0,175 0,202 0,295 0,225 0,283 0,295 0,346

7,8 7,5 6,8 3,5 6,2 3,1

8,0 4,8 3,0 6,3

5,2 4,9 4,0 4,2 6,6 4,0 5,9 6,4 7,4

0,718--0,998 0,842--1,112 0,936--1,373 1,123--1,279 1,148--1,560 1,373--1,560 1,404--1,560 1,404--1,560 1,498--2,059 1,934--2,496 2,402--2,683 2,184--2,870

2,714--3,588 3,048--3,861 3,708--4,775 3,810--4,724 4,429--5,080 3,658--5,080 4,572--5,385 4,369--5,385 3,861--5,131 3,810--5,283

8*

100 64 64 36 36 32

5 13 36 36 36 19

1 24 35 33 33 33 35 34 36 36 36

o ~

c~

Q

c~

H ~

I~

Q~

CD

N ~


Table7. Size of the pupae.

M • a = • C

/ Chubby . . . . Leng*h Wild-type . . .

Width ~ Chubby . . . . Wild-type . . . (

2,521 • 2,945•

1,154={=0,008 1,064•

0,135 0,166

0,077 0,074

5,4 5,6

6,7 7,0

Lim

2,235--2,845 2,591--3,302

1,016--1,422 0,914--1,219

100 100

100 100

and pupae. I t remains to be de termined whether the rate of growth of the c h u b b y larvae is the same as t ha t of wild-type, Since the ini t ia l dimensions of the larvae are dissimilar, the dissimilari ty of the advanced

400 . < r . . . . . . o ......

. . .o . . . . o - . ,

3.00 ...... " ~ .......

I.so l l 6

lO0 o/''g' Cc80

d l 0.7o I I I I f ] I T t I P I I I I k f

4 8 12 16 20 24 30 36 42 48 54 60 66 72 78 84 90 96

Fig. 3. Growth of the length of the body in chubby and wild-type. Abscissae -- age in hours; ordinates -- body length; solid line -- wild type; dotted line -- chubby. Roman

figures indicate the instars.

stages of development m a y be due to mere persistence of the ini t ia l difference. On the other hand, the ini t ia l difference ma y be augmented or decreased if the growth rates are different.

The results of the measurements of the larvae of different ages are shown in Tables 3- -6 . The same results are represented graphical ly in Figs. 3 and 4. The measurements of the pupae are given in Table 7. Growth proceeds a t a ra ther un i formly decreasing rate from the emergence of the larvae from. the eggs up to and including the beginning of the th i rd instar. I t slows down rapid ly in the th i rd instar , a nd ceases a t abou t the age of 78 hours. The growth is apparen t ly discontinued, or even a decrease of the size of the larvae is t ak ing place, a t the end of each instar. This is, probably, due to the overlaping of the instars, as poin ted


out also by AgPATOV (1929). Some larvae undergo moulting while others continue to feed. I t is not improbable that those larvae which continue feeding while their sibs arc undergoing moulting or pupation arc somewhat handicaped in their development by some external or internal factors. This may also account for the apparent sudden increase of the size of the larvae observed a t the transition from one instar to the next one (see, for example, the increase of chubby between the second and the third instar).

I.m / ..~ ,..~-.--o..,_ -~--

I . t 0 " ' ' ' - 0 ' ' "

Qao .... ~"

O~ , too'' "

04o m / : ' / ( "

O~ ,.", " I . . . . . .

,/ Q'~

, , , , , , , , , , , 1 4 8 12 162024 30 3G 42 48 ,54 60 6G 72 78 84 90 96

Flg. 4. Growth of t h e ~ i d t h o f t h e b o d y i n chubby and wild-type. ~bsclssae -- a g e i n hours; ordinates -- body width ; solid line -- ~vild tYPe; dotted line -- chubby.

The growth curves for chubby and wild-type on the whole run parallel to each other . . Only in the third instar do the growth curves for the length of the body clearly diverge, and those for the width of the body converge. Chubby and wild-type grow, therefore, equally fast up to the end of the second instar, while in the third instar the growth of chubby is somewhat slowed down. As shown above, this slowing down of the growth of chubby is reflected also in the body weight. This fact does not decrease, however, the difference between chubby and wild-type, since the relative difference (i. e. the ratio between the length and the width of the body) remains fairly constant.

A more precise comparison of the growth rates can be made by calculating the growth coefficients (C I ) . According to SC~AT.~'AUSE~T (1928, 1930 a, b), the growth coefficient is calculated from the formula:

log L1 - - log L C1 - - 0,4343 (t~ - - t )

where L and L x are the lengths of the body at times t and t 1 respectively. The growth coefficients for chubby and wild-type are shown in Table 8.


Table 8. Growth c o e f f i c i e n t (C1) for c h u b b y and w i l d - t y p e (pro one hour).

I I

I I I

Time interval

1--4 4--8

I 8--12 12--16 16--2o 20--24 24---30 30--36 36 -42 ~ - - 4 8 48--54 54---60 60--66 66--72 72--78

400

500

Length of the bo4y

Chubby Wild-type

0,0565 0,0543 0,0299 0,0332 0,0058 0,0169 0,0329 0,0224 0,0246 0,0296 0,0202 0,0328 0,0274 0,0386 0,0177 0,0231 0,0162 0,0055 0,0184 0,0179 0,0141 0,0385 0,0193 0,0049 0,0031 0,0122 0,0070 0,0041 } 0,0025

Widbh of the body

Chubby

0,0563 0,0303

0,0127

0,0223 0,0175 0,0207 0,0325 0,0171 0,0162 0,0270 0,0110 0,0041

0,0139

Wild-type

0,0272 0,0126 o, o12o 0,0515 0,0260 0,0363 0,0281 0,0191

0,0156

0,0104 0,0223

0,0047

4

Y

[,o;{; ...........

96 9Q

.,,'1~5 4 y-"

,..u48

a'42

/

"~ ....."

/

0.~ /

0.7r c I 0.17 020 030 0.40 0.60 0.80 to0

Fig. 5. /~ double logarithmic plot of the body length (ordinates) against bogy-width (abscissae). Solid line -- wild-type; dotted line -- chubby. Figures indicate age in hours.

The g rowth coefficients tend , on t h e whole, to decrease wi th age, especial ly in the t h i rd ins tar . The process proceeds in para l le l manner in c h u b b y and wi ld- type . The values of the growth coefficient for s imilar t ime in tervals a re somet imes higher for c h u b b y t h a n for wi ld - type and


sometimes the reverse relation is observed. There is no pronounced tendenncy for the growth of the length of the body to be faster in wild-type than in chubby, nor for the width of the body to be faster in chubby than in wild-type. The larvae of the two types grow at approximately equal rates. The difference observed between the mature larvae of chubby and wild-type is, consequently, to be ascribed to the persistence of the initial difference present a t the moment of hatching from the egg. Neither an exaggeration nor an inhibition of the initial difference due to a dissimilarity of the growth rates is observed.

A further check of this conclusion is afforded by studying the degree of heterogony manifested in the larval growth in chubby and ~ l d - t y p e (comp. HuxLEY 1932). A double-logarithmic plot of the length of the body against the width of the body in chubby and wild-type larvae is shown in Fig. 5. The curves obtained approximate straight lines, anc[ are parallel to each other. The growth of chubby and wild-type obeys the same law.

Absence of maternal effect. The rate of growth of chubby and wild-type larvae is alike. One may

conclude tha t the difference between chubby and wild-type is due to an alteration of the course of development produced by the gene chubby before the ecelosion of the larva from the egg. This conclusion may be restated as follows: there is no evidence that, as far as the shape of the body is concerned, the difference between the genes chubby and its wild-type allelomorph influences the development during the larval stage 1.

At what stage of the development, then, does the gene chubby act ? I t must act before the larva leaves the egg shell, but it remains to be determined how long before this moment the difference between chubby and wild-type is produced. I t may happen during embryonic development, or even before fertilization and before the egg leaves the mother 's body. The genetic constitution of the mother may influence the egg developing in the ovary, and this influence may determine its behavior after fertilization. The fact tha t the egg shape is alike in chubby and wild-type (see above) certainly does not exclude such a possibility. Indeed, the cytoplasm of the egg rather than its chorion might be influenced.

The possibility of the effect of chubby before fertilization can be tested experimentally. For, if the behavior of the egg is determined before fertilization, a maternal effect eou]d be observed. Individuals

1 The reservation "as far as the shape of the body is concerned" is an important one, for we do not know the total effects of the genes chubby and its wild-type allelomorph, nor do we know whether the difference between these two genes influences only the shape of the body, or also some other characteristics which escape our notice.

Genes tha~ affect early developmental stages of Drosophila melanogaster. 121

Table9. Length of the body in larvae coming from chubby eggs.

~ .ge i n h o u r s 3I • m a = -4- C L i r a n

I I

Newborn 3 5 7 9

12 16 2O

III

20 24 30 36 42 54 66 78

0,916=[=0,009 0,986• 1,123• 1,191• 1,231• 1,398~0,017 1,534• 1,509!0,021 2,035J=0,060 1,965~0,033 2,711 =t=0,022 2,839-4-0,018 3,450::[:0,04-5 4,629:t::0,047 5,104q-0,049 4,917•

0,073 0,067 0,072 0,071 0,146 0,083 0,076 0,078 0,272 0,165 0,112 0,092 0,216 0,237 0,247 0,228

7,9 6,8 6,4 5,9

11,8 5,9 5,0 5,2

13,4 8,4 4,2 3,8 6,3 5,1 4,8 4,6

0,660--1,067 0,813--1,067 1,016--1,270 1,118--1,372 1,016--1,626 1,270--1,575 1,372--1,676 1,422--1,676 1,626--2,591 1,778--2,591 2,489--2,997 2,591---3,048 3,048--3,912 4,166--5,182 4,572--5,436 4,572--5,486

72 32 36 25 25 25 25 14 20 25 25 25 25 25 25 25

Table 10. Length of

A g e i n h o u r s

Newborn 3 5

I 7 9

12 16 2O 20

I I 24 3O 36 42

III 54 66 78

the body in larvae coming

l~I ~ rn a = ~ = C

0,897-4-0,009 1,047-4-0,012 1,082-4-0,014 1,177• 1,252• 1,445• 1,477• 1,504-4-0,038 1,807• 2,146~=0,054 2,715• 2,898• 3,552=}=0,045 4,275• 5,096• 4,905•

0,068 7,5 0,065 6,2 0,075 6,9 0,083 7,0 0,082 6,6 0,079 5,5 0,092 6,2 0,121 8,0 0,110 6,1 0,273 12,7 0,197 7,3 0,100 3,4 0,227 6,4 0,183 4,3 0,194 3,8 0,240 4,9

from wi ld . type eggs.

L i m n

0,762--1,016 52 0,864--1,168 31 0,914--1,168 27 1,016--1,422 25 1,118--1,422 25 1,270--1,626 25 1,270--1,626 25 1,270--1,676 10 1,626--2,032 23 1,727--2,845 25 2,134--2,946 25 2,642--3,150 25 3,048--4,064 25 3,810--4,572 25 4,623--5,486 25 4,420--5,537 25

having identical genic constitution but coming from different kinds of eggs must be different. Crosses of chubby females with wild-type males, and of wild-type females with chubby males were made. Since chubby is an autosomal recessive, the offspring of either of these crosses consists of individuals heterozygous for chubby (chubby/A-). If, however, chubby acts before fertilization the offspring coming from chubby eggs must be different from that coming from wild-type eggs.

The growth of the larvae coming from chubby eggs, and of those coming from wild-type eggs, was studied. The results are shown in Tables 9--12. Little comment is necessary for ~hese results. There is no difference between larvae coming from chubby and from wild-type eggs, provided the genetic constitution of these larvae is identical. The


Table ll. Width of the body larvae coming from chubby eggs.

II

I I I

Age In hours

Newborn 3 5

I 7 9

12 16 2O 2O 24 3O 36 42 54 66 78

0,193::[:0,001 0,240-+-0,003 0,276:t:0,005 0,268-+-0,006 0,289-4-0,008 0,307• 0,345-}-0,007 0,330 - - 0,386+0,012 0,354-4-0,007 0,549-4-0,008 0,567 :]::0,0O7 0,627• 0,872+0,014 0,994-4-0,012 1,038-4-0,018

a=-t-

0,012 0,018 0,031 0,029 0,040 0,030 0,034

o,~5 0,034 0,038 0,037 0,041 0,072 0,061 0,092

C

6,1 7,2

11,2 10,7 13,8 10,0 9,5

1 ,2 9,6 6,8 6,6 6,6 8,1 6,2 8,9

Ltm

0,152--0,229 0,203--0,254 0,203--0,356 0,254--0,356 0,254--0,406 0,254--0,356 0,305--0,406

0,305--0,457 0,305--0,457 0,457--0,610 0,508--0,610 0,508--0,711 0,711--1,016 0,864--1,118 0,864---1,219

72 32 36 25 25 25 25 3

20 25 25 25 25 25 25 25

Table 12. Width of

A.ge in hours

Newborn 3 5

I 7 9

12 16 20 2O 24

II 30 36 42

III 54 66 78

the body in larvae coming from wild-type eggs.

1~ :i: m

0,199• 0,250-4-0,002 0,273-4-0,006 0,274-4-0,006 0,266+0,004 0,305t::0,006 0,323:l:0,0O8 0,305-4_-0,009 0,338:[:0,003 0,378!:0,011 0,538• 0,561 i0,008 0,652+0,008 0,792-+-0,010 0,996• 1,006::[:0,014

a=-t-

0,014 0,014 0,028 0,029 0,021 0,029 0,040 0,023 0,015 0,053 0,041 0,039 0,034 0,052 0,069 0,072

1 1 8,1 9,5

12,4 7,5 ~,6

1 ~,2 7,5 7,0 5,2 6,6 6,9 7,1

c Lim

7,3 0,178---0,254 5,8 0,203---0,254 D,4 0,203---0,356 ~),5 0,254---0,356

0,254---0,305 0,254--0,356 0,254---0,356 0,254--0,381 0,305---0,406 0,305--0,508 0,406---0,610 0,508---0,610 0,533---0,711 0,686--0,914 0,813--1,118 0,864--1,143

52 31 27 25 25 25 25 10 23 25 25 25 25 25 25 25

gone chubby produces no maternal effect. The same conclusion follows from the fact that the larvae appearing in F~ generation from the cross chubby X wild-type can be easily classified into chubby and wild-type; this would hardly be the case if chubby produced a strong maternal effect. There is, hence, no evidence that the gone chubby acts before fertilization, at least as far as the shape of the larvae is concerned. The action of the gone chubby seems to be restricted to the period of the embryonic development only.

Cases of maternal effect are rare in Drosophila. Mo~G~-~ (1915) and LY~c~ (1919) found that the presence in the eggs of certain genes before fertilization may make these eggs incapable of development if fertilized by spermatozoa carrying these genes. RED~LD (1926) described


a sex-limited lethal effect inherited maternally. HERS~ (1927) found that the number of eye-facets in the mutant Bar is subject to maternal effect (Hv.Rs~s conclusion is, in the opinion of the writers, not sufficiently well established). The scarcity of the known cases of maternal effect in Drosophila is not surprising. I t is, probably, due to our ignorance concerning genes affecting larval characters. In insects with necrobiotic metamorphosis, to which Drosophila belongs, the body is nearly com- pletely rebuilt in the pupal stage. The chance of finding adult characteristics exhibiting maternal effects is, therefore, rather small.

On the nature of the chubby character. An at tempt was made to determine which parts of the body are most

strongly affected by the gone chubby. The short and stout appearance of the chubby larvae and pupae might be due to the integument and the muscles alone, or to a decrease in length and an increase in width of all internal organs as well. Mature larvae (72 hours old) were dissected, and their internal organs were studied. The nerve ganglia and the gonads seem to be similar in chubby and in wild-type. The alimentary canal was isolated and drawn with aid of a camera lucida. The length of the alimentary canal (from the pyloric glands to the 2M'AT~T'XGm_A~ vessels) was measured in twenty-five chubby and in an equal number of wild- type larvae 1. The mean length of the intestine was determined to be (in mm):

chubby . . . . . . . 6,14 -V 0,22 wild-type . . . . . . . 5,90 ~: 0~20.

There is, evidently, no significant difference between chubby and wild-type in this respect. I t seems that chubby does not affect the shape of the organs lying in the body cavity. The most probable hypothesis is that the main effect of chubby is on the larval musculature. I f one observes the movements of the anterior end of the larvae (including the mouth-armature), it appears that the chubby larvae are much less able to extend and to retract these parts than the wild-type. The movements of the chubby larvae seems to be rather more sluggish. This hypothesis can, of course, be proven only by a detailed morphological study.

I t may be emphasized here tha t the adult chubby and wild-type flies are far less strikingly different from each other than are their larvae and pupae. This fact may be correlated with the extreme violence of the processes taking place in the pupa, Not only is the larval hypoderm and musculature destroyed but most of the internal organisation of the larva as well, and the organism is rebuilt from the imaginal disc. I t

i The drawings of the alimentary canal were measured with aid of a thin copper wire which could follow all the curvatures of the gut. This method of measurement was suggested by Mr. W. ZA~XOWSK~, whose help the writers wish to acknowledge.

19,4 Th. Dobzhansky and F. ~N. Duncan:

appears, then, tha t the imaginal discs can construct an approximately normal organism even in case when their spatial relations are as different as they must be in pupae of chubby and wild-type. Here we seem to have an instance of a fairly independent evolution of the larval and the adult stages, a problem rather frequently discussed by insect morpho- logists.

Growth in rudimentary-12 larvae. As pointed out above, the difference in the shape of the body of

larvae and pupae observed between rudimentary-12 and wild-type may be due either to the effect of the gene-rudimentary-12 itself, or to the effect of an independent factor closely linked with rudimentary. This makes the situation observed in rudimentary-12 less easy to interpret than the situation encountered in chubby. I t remains, indeed, possible tha t the differences observed between rudimentary-12 and wild-type are due to an interaction of two independent loci.

As far as our data go, the effect of rudimentary-12 on the shape of the larvae and pupae is highly analogous to the effect of chubby on the same character. Such a similarity of the effects of different genes located in different chromosomes is not rare in Drosophila. For instance, the genes vermilion (X-chromosome), cinnabar (second chromosome), scarlet and cardinal (third chromosome) produce very similar changes of the eye color. The effects of chubby and rudimentary-12 on larvae and pupae are not allelomorphic. The offspring of the crosses chubby 9 • rudimentary-12 ~ and rudimentary-12 9 X chubby 3 consists of wild-type larvae.

Table l3. Length of the body of rud imenta ry-12 larvae.

.Age i n hours

I I

I I l

Newborn 6

I 12 18 24 24 30 36 42 48 54 60 66 72 78 84 90 96

�9 I :t= m

0,72110,007 0,821i0,008 1,029~0,015 1,124• 1,235=L0,020 1,501t0,091 1,673+0,024 2,042~=0,017 2,091!0,020 1,946• 3,017=j=0,045 3,235=t=0,050 3,293• 3,481i0,019 3,462-4-0,032 3,443=3=0,051 3,374• 3,479•

a = •

0,069 0,041 0,076 0,103 0,077 0,242 0,122 0,084 0,116 0,128 0,222 0,239 0,214 0,166 0,204 0,267 0,144 0,200

9,6 5,0 7,4 9,2 6,3

16,0 7,3 4,1 5,5 6,6 7,4 7,4 6,5 4,8 5,9 7,7 4,3 5,7

Lim

0,506---0,853 0,758--0,916 0,916--1,201 0,727--1,264 1,106--1,454 1,264--1,959 1,517--1,801 1,864--2,244 1,865--2,279 1,709--2,176 2,486--3,419 2,486--3,471 2,901--3,522 3,004--3,730 3,160--3,885 2,590--3,885 2,901--3,626 3,056--3,885

n

100 25 28 26 15 10 25 25 30 28 24 23 24 27 27 27 27 24


Table 14.

2~ge in hours

Newborn 6

12 I 18

24 30 36 24 30 36

I I 42 48 60 48 54 60 66

I I I 72 78 84 9O 96

L e n g t h of

:M:•

0,767=t=0,009 0,922• 1,129=J=0,029 1,273i0,025 1,465• 1,571-t-0,077 1,454 - - 1,675 - - 1,886=]=0,028 2,367-t-0,023 2,497=]=0,023 2,721-4-0,030 2,694 - - 3,060• 3,461 • 3,700• 4,172• 4,.618i0,050 4,536~0,041 4,566-t-0,055 4,666=t=0,041 4,381•

the body of w i l d - t y p e larvae. a = •

0,090 0,035 0,104 0,126 0,070 0,109

0,130 0,112 0,126 0,138

0,329 0,257 0,270 0,292 0,271 0,224 0,292 0,217 0,260

c Lira

11,7 0,569--1,011 3,8 0,790--0,980 9,2 0,853--1,327 9,9 0,916--1,422 4,7 1,296--1,580 6,9 1,454--1,770 - - 1,454 - - - - 1,548--1,864 6,9 1,548--2,149 4,7 2,117--2,528 5,0 2,227--2,745 5,1 2,538--3,108 - - 2,694 - - 0,8 2,435--3,626 7,4 2,745--3,937 7,3 3,004---4,248 7,0 3,522--4,662 5,9 3,937--5,128 4,9 3,937---4,869 6,4 4,144--5,128 4,6 4,196--5,025 6,0 3,730--4,869

100 24 27 25 23 55

1 3

22 24 31 22 2

14 30 25 27 29 29 28 28 26

Table l5. W i d t h of the body of r u d i m e n t a r y - 1 2 larvae .

Age in hours

Newborn 6

I 12 18 24 24 30

I I 36 42 48 54 6O 66

I I I 72 78 84 90 96

~Vi•

0,191~0,002 0,195• 0,253~0,005 0,288i0,007 0,310i0,007 0,348i0,024 0,386-4-0,006 0,478=t=0,013 0,520~0,010 0,470• 0,75110,015 0,759q-0,019 0,846• 0,929• 1,005:J:0,013 1,007• 0,907:J:0,012 0,984i0,013

a = •

0,022 0,019 0,025 0,036 0,028 0,058 0,031 0,064 0,057 0,062 0,072 0,090 0,074 0,101 0,067 0,095 0,061 0,065

C

11,7 9,9

10,4 12,7 9,0

16,6 8,2

13,2 11,1 13,1 9,6

11,9 8,8

10,8 6,6 9,4 6,7 6,6

Lira

0,158--0,253 0,158--0,221 0,221--0,316 0,158---0,348 0,253--0,379 0,284---0,474 0,316--D,411 0,316--0,537 0,414--0,622 0,363--0,570 0,622---0,881 0,518--0,932 0,673--1,036 0,570--1,036 0,829--1,088 0,622--1,140 0,777--0,984 0,777--1,082

100 25 29 26 16 9

25 25 30 28 24 23 24 27 27 27 27 24

Tables 13- -16 summarize the results of the measurements of the larvae of rud imentary-12 a n d wild-type 1. The newborn larvae of rudi-

1 These wild-type larvae were raised and fixed simultaneously with the rudimentary-12 ones. A comparison of the data for wild-type larvae presented in Tables 14 and 16 with those in Tables 4 and 6 shows that these two sets of data are somewhat different. This fact shows that different kinds of larvae which are to be compared with each other must always be reared simultaneously, under identical temperature and food conditions.


Table 16. W i d t h of the body of w i l d - t y p e - l a r v a e .

A g e in hou r s

Newborn 6

12 18 24 3O 36 24 3O 36 42 48 6O 48 54 6O 66 72 78 84 90 96

~ •

0,182• 0,178-4-0,003 0,211 ~0,006 0,235-4-0,006 0,284~0,003 0,284=t=0,034 0,284 - - 0,326 - - 0,319• 0,402• 0,444~0,007 0,499• 0,466 - - 0,529=]=0,020 0,570=]=0,012 0,597• 0,700• 0,813=t=0,019 0,813-4-0,011 0,879=]=0,018 0,914• 0,912•

0,019 0,015 0,030 0,032 0,013 0,034

0,061 0,036 0,039 0,065

0,065 0,064 0,066 0,067 0,105 0,058 0,093 0,044 0,069

10,9 8,6

14,1 13,7 4,6

12,2

19,2i 8,9 7,8

13,1

1 2 , 3 1],2 i 11,9 9,5

12,9 7,1

10,6 4,8 7,6

Lim

0,158~0,253 0,158--0,190 0,158--0,253 0,158-0,348 0,253-0,316 0,221-0,316 0,284 - - 0,316-0,348 0,253---0,379 0,348-0,474 0,311-0,578 0,414-0,673 0,466 - - 0,414--0,622 0,466-0,673 0,466--0,673 0,570-0,777 0,570--1,036 0,673-0,932 0,725--1,036 0,829--1,036 0,725--1,036

n

100 24 27 26 23

5 1 3

23 24 31 22 2

14 30 25 27 30 29 28 28 26

5.C

4~

ZC -

I.~ "n'o / .o" ....

0.8 . . . .

I 6 12 18 24 30 36 42 48 54 60 66 72 78 84 90 96 Fig. 6. Growth of the length of the body in rudimentary-12 and wild-type. ~kbscissae - - age in hours; ordinates -- body length; solid line -- wild-type; dotted line -- rudimentary-12.

mentary-12 are a l ready clearly different f rom wild-type. The former are 0,046 • 0,011 m m shorter but 0,009 • 0,003 mm wider t han the lat ter . A t the age of 6 hours the differences are ful ly significant f rom the stati-

stical viewpoint .

Genes that affect early developmental stages of Drosophila melanogaster. 19, 7

The growth curves for rudimentary-12 and wild-type (Figs. 6 and 7) are rather closely similar in the first and the second instars. In the third instar rudimentary-12 larvae become rather smaller than wild-type larvae of similar age. Consequently, the curves for the body length

0.6c

0.4c

O.,~c

O.ac

0.17 I I I I I I I 6 12 18 24 30 36 42 4.8 54 60 66 72 78 84 90 96

Fig. 7. GroWth of the width of the body in rudimentary-12 and wild-type. 2kbscissae - - age in hours; ordinates -- body width; solid line -- wild-type; dotted line -- rudimentary-12.

diverge, and those for the body width converge in the third instar. A similar phenomenon was encountered in comparing chubby and wild-type (see above), but, here it is more pronounced. In both cases, however, only the absolut~ dimensions are affected, the ratio between the length and t'he width of the body remaining more or less constant.

Discuss ion .

Among the various possible types of developmental changes two types seem to be especially interesting. These are, first, the early embryonic changes, and, second, changes due to an early termination or extension of development. The former, if not regulated in later development, al ter the appearance of the organism from the embryonic to the adult stage. The latter leave most of the embryogeny except the latest stages unaffected. The possibility of these two types of changes was discussed more than 100 years ago by the great German-Russian embryologist vow BAER (1828).

Cases of chubby and, possibly, of rudimentary-12 are examples of the early developmental variations. These genes cause an alteration of the embryonic development. Larvae leaving the egg shells are different. This difference is merely preserved, but not accentuated, during the larval and the pupal development. During the reconstruction of the

128 Th. Dobzhansky and 1~. N. Duncan:

body in the pupa a regulation process takes place, and the adult chubby and wild-type are rather similar in appearance. The very interesting work of D ~ R I c x (1928), unfortunately published only in a very pre- liminalT form, indicates that the development of the mutant eyeless proceeds in a manner similar to that observed in chubby. Dr. N. M~.D- vrD~v, of the University of Leningrad, also informs us that according to his unpublished data ~he differences between various Drosophila mutants affecting eye-shape are noticeable in early stagcs of the development of the imaginal discs. Similar phenomena are probably observed in the development of mutants affecting wing size in Drosophila (CHE~ 1929).

An example of a change due to an extension of development is the mutant giant in Drosophila melanogaster (GABRITSCts165 and BRID- GES 1928). The larvae of giant are, as far as known, not distinguishable from their normal sibs in the early stages. But they continue feeding and their growth is extended for a considerably longer period than is the case in normal larvae. I t seems possible that giant has four instead of three larval instars, but this question has not been studied.

Which of the two types of the developmental changes is more frequent among the Drosophila mutants and elsewhere can be decided or ly by future investigations. This problem is of significance both for genetics and for the theory of evolution and warrants more attention from investi- gators than it has received heretofore.

Summary. 1. The second -chromosome recessive, chubby, affects the body shape

of larvae, pupae, and, to a lesser extent, of the adult. Chubby larvae are shorter but broader than wild-type. The sex-linked recessive rudimentary-12, or a factor closely linked with it, affects larvae and pupae in a manner similar to chubby.

2. The weight of chubby and wild-type larvae and pupae is nearly alike. 3. The shape of chubby eggs is similar to wild-type. 4. The shape of the body in the newborn chubby and rudimentary-12

larvae is clearly different from wild-type. 5. The rate of growth of chubby and rudimentary-12 larvae is similar

to wild-type. The difference in shape, already present at the emergence of larvae from the eggs, is merely preserved during the larval and the pupal development.

6. Chubby produces no maternal effect. 7. The effect of chubby on the body shape is produced between the

time of fertilization and emergence of larva from the egg. 8. Chubby does not affect the shape of the internal organs of larvae.

Its effect seems to be restricted to the larval musculature, and, probably, the hypoderm.

Genes that affect early developmental stages of Drosophila melanogaster. 19,9

Zusammenfassung.

1. Die K~rper fo rm yon La rven und Puppen und in ger ingerem Mal~e yon erwachsenen F l iegen wird durch das rezessive Gen , , chubby" beein- fluBt. Das Gen c h u b b y ist in dem zweiten Chromosom lokal is ier t . Chubby-Larven und -Puppen sind merkl ich kfirzer und p lumper als bei der Wi ldform. Eine ziemlich gleiche Modif ikat ion der KSrper fo rm yon La rven und P u p p e n wird auch durch das geschlechtsgebundene Gen , , rud imentary-12" , oder du t ch ein mi t ihm eng gekoppel tes Gen produzier t .

2. Die KSrpergewichte von La rven und P u p p e n s ind bei , , chubby" und Wfldform ziemlich gleich.

3. Die E ie r yon chubby untersche iden sich n icht yon den Eie rn der Wildform.

4. Die KSrper fo rm der soeben aus den E ie rn ausgeschlf ipften La rven ist bei , , chubby" u n d , , rud imenta ry-12" deut l ich verschieden yon der Wildform.

5. Die Wachs tumsgeschwind igke i t is t sowohl fiir c h u b b y als aueh fiir r ud imen ta ry -12 -La rven gleich der des Norma l typus . Die Differenz in der Fo rm, die schon be im Ausschli ipfen aus dem Ei in Ersche inung t r i t t , b le ib t im Larven- und P u p p e n s t a d i u m erhal ten, und wird weder st/~rker noch schw/icher.

6. Chubby ruff keinen materne l len Effekt hervor . 7. Der EinfluB von chubby auf die KSrper fo rm f indet zwischen der

Befruchtung und dem Ausschlt ipfen der Larve s tar t . 8. ] )as Gen chubby ha t keinen EinfluI~ auf die Ges ta l t der inneren

Organe der Larven. Sein EiIffluB scheint auf die Musku la tu r und wahr- scheinlich auf die Hypode rmi s der La rven beschr~nkt zu sein.

Literature cited. Alpatov, W. W.: Growth and variation of the larvae of Drosophila mslanogaster.

J. exper. Zool. 52, 402--437 (1929). - - Growth of larvae in wild Drosophila melanogaster and its mutant vestigial. J. exper. Zool. 56, 63--71 (1930). - - Baer, K. E. yon: l~ber Entwicklungsgeschichte der Tiere, Bd. 1. K6nigsberg: Borntr/~ger 1828. - - Bridges, C. B.: Non-disjunction as proof of the chromosome theory of heredity. Genetics 1, 1--52, 107--163 (1916). - - Chen~ T. Y.: The development of imaginal buds in normal and mutant Drosophila melanogaster. J. Morph. a. Physiol. 47, 135--199 (1929). - - Derrick, G. E.: The development of the eye and optic tract in Drosophila melanogaster and its "eyeless" mutant. Univ. Oklahoma Bull. 8, 100--105 (1928). ~ Dobzhansky, Th.: Studies on the manifold effect of certain genes in Drosophila melanogaster. Z. Abstammgslehre 43, 330--388 (1927). - - Gabritsehevsky, E. und C. B. Bridges: Physiological aspects of the giant race. Z. Abstammgslehre 46, 248--284 (1928). - - Hersh, A. H.: Temperature effects in reciprocal crosses of the Bar series of Drosophila. J. expel Zool. 47, 227--250 (1927). - - Huxley, J. S.: Problems of relative growth. The Dial Press. New York 1932. - - Li, Y. C.: The effect of chromosome aberrations on development in Drosophila melanogaster. Genetics 12, 1--58 (1927). - - Lynch, C. J.: An analysis of certain eases of intra- specific sterility. Genetics 4, 501--533 (1919). - - Mohr, 0. L : Cases of mimic

W. l~oux' 2~rchiv f. Entwicklungsmeehanik. Bd. 130. 9

180 Th. Dobzhansky and F. N. Duncan.

mutations and secondary mutations in the X-chromosome of Drosophila melanogaster. Z. Abstammgslehre 28, 1--22 (1922). - - Morgan, T. H.: The infertility of rudimentary winged females of Drosophila ampdophila. Amer. Naturalist 49, 240--250 (1915). - - The scientific basis of evolution. New York: Norton & Co. 1932. - - Morgan, T .H. , C.B. Bridges and A. H. Sturtevant: The genetics of Drosophila. Bibliogr. Genetics 2, 1--262 (1925). - - Philiptehenko, J . : Gene und Entwieklung der ~hrenform beim Weizen. Biol. Zb]. 49,1--16 (1929). - - Red|ield, H.: The maternal inheritance of a sex-limited lethal effect in Drosophila melanogaster. Genetics 11, 240--250 ( 1 9 2 6 ) . - Sehmalhausen, L: Das Waehstumsgesetz und die Methode der Bestimmung der Wachstumskonstante. Arch. Entw.mechan. 113, 462--519 (1928). - - ~ber Wachstumsformeln und Wachstumstheorien. Biol. Zbl. 50, 292--307 (1930a). - - Das Wachstumsgesetz sis Gesetz der progressiven Differenzierung. Arch. Entw.- mechan. 123, 153--178 (1930b). - - Srhultz, J. : The minute reaction in the development of Drosophila melanogazter. Genetics 14, 366--419 (1929). - - Sivertzev- Dobzhansky, N. P.: t?ber den letalen Effekt einiger Gene bei Drosophila melanogaster. Arch. Entw.mechan. 109, 535--548 (1927). - - Stark, M. B.: An hereditary tumor in the fruit fly, Drosophila. J. Cane. Res. 8, 279--301 (1918). - - A benign tumor that is hereditary in Drosophila. Prec. nat. Aead. Sci. U.S.A. 5, 573--580 (1919). - - Stark, M. B. and C. B. Bridges: The linkage relations of a benign tumor in Droso- phila. Genetics 11, 249--266 (1926). - - Warren, D. C.: Inheritance of egg size in Drosophila melanogaster. Genetics 9, 41--69 (1924).

genes that affect early developmental stages of drosophila melanogaster

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