Download Cytogenetic Analysis Shows that the Unusually Large Chromosome

Hereditas 133: 95- 103 (2000)
Cytogenetic analysis shows that the unusually large chromosome in
the sex-limited p B silkworm (Bombyx mori) strain consists of three
chromosomes
NOBUHIKO TANAKA' , TAKESHI YOKOYAMA', HIROAKI ABE', KOZO TSUCHIDA',
OSAMU NINAGI' and TOSHIKAZU OSHIKI'
Department of Biological Production, Faculty o j Agriculture, Tokyo University of Agriculture and
Technology, Saiwai-cho 3-5-8, Fuchu, Tokyo 183-8509, Japan
National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku-ku, Tokyo 162-8640, Japan
Tanaka, N., Yokoyama, T., Abe, H., Tsuchida, K., Ninagi, 0. and Oshiki, T. 2000. Cytogenetic analysis shows that the
unusually large chromosome in the sex-limited p B silkworm (Bombyx mori) strain consists of three chromosomes.
Hereditas 133: 95-103. Lund, Sweden. ISSN 0018-0661. Received August 7, 2000. Accepted October 30, 2000
~
We have discovered an inordinately large chromosome pair at the pachytene stage in the oocyte of the sex-limited p B
(black larval marking) silkworm (Bombyx mori) strain (TWPB). We have analyzed the composition and arrangement of
this large chromosome. A genetic linkage analysis shows that the large chromosome is made up of the W chromosome,
the second chromosome fragment (p fragment), and the fifth chromosome (linkage group) containing at least the region
from map position 0.0 to 40.8. We also observed a sex heterochromatin body (SB) that we deduced to be made up of
condensed W chromosomes. The number of SBs in each female nucleus among the sucking stomach cells of the TWPB
strain was variable. Evidently, the W chromosome of the TWPB strain is attached to another chromosome. The
composition of the W chromosome, the second chromosome fragment, and the fifth chromosome was studied through
linkage analysis for these three chromosomes. We used two strains derived from the TWPB strain, the sex-limited p M
(moricaud barVal marking)-hke (TWPML) and the autosomal p M-like (TSPML). The results show that the TWPML strain
originates through a detachment of the fifth chromosome from the large chromosome of the TWPB strain, and the
TSPML strain originates through a detachment of the W chromosome from that. Accordingly, the large chromosome of
the TWPB strain is arranged in the order W chromosome-second chromosome fragment-fifth chromosome.
Nobuhiko Tanaka, Laboratory of Sericulturul Science, Depariment of Biological Production, Faculty of Agriculture, Tokyo
University of Agriculture and Technology, Saiwai-cho 3 - 5 8 , Fuchu, Tokyo 183-8509, Japun. E-mail: [email protected]
TANAKA(19 16) showed that females of the silkworm,
Bombyx mori, are ZW, and males are ZZ. The female
sex is determined through a single W chromosome.
This was confirmed by the observation of polyploids
(HASIMOTO1933; KAWAGUCH~
1934), a sex chromosome non-disjunction strain (TANAKA1939), and sexlimited strains (TAZIMA1944).
Chromosomal constitutive aberrations such as
translocation, duplication, deletion, and attachment
have been frequently found in B. mori (TAKASAKI
1952; FUJIIet al. 1998). Strains that have an autosoma1 fragment having a dominant gene for egg color,
larval marking, and blood color translocated on the
W chromosome are useful because sex can easily be
determined through the sex-limited expression of
characteristics (TAZIMA1941; TAZIMAet al. 1951;
HASIMOTO1948; KIMURAet al. 1971). TAZIMA
(1941) found the sex-limited p s u (sable larval marking) strain. This strain had a translocation between
the W and the second chromosome having a mutant
allele in the p locus, which controls larval markings.
TAZIMA(1942, 1943, 1944) irradiated this sex-limited
strain with X rays and obtained a sex-limited normal
marking ( + ") strain, which had eliminated the exces-
sive part of the translocated second chromosome.
The following strains originated spontaneously within
these strains: ones with the translocated W chromosome, derived from the sex-limited + P (normal larval
marking), sex-limited p (moricaud larval marking)
(TAZIMAet al. 1955), sex-limited p B (black larval
marking) (SASAKI1955), and other sex-limited
strains (SASAKI1958).
The chromosomes of B. mori are small, many
(2n = 56), and not available to banding techniques, so
that each individual chromosome has not yet been
cytologically identified. Spermatocytes have been in
general used for observing chromosomes because it is
relatively easy to obtain chromosome images in dividing cells. Chromosomes at the pachytene stage from
the oocyte are more suitable for observing chromosome morphology, even though it is very difficult to
make good chromosome preparations in the female
pachytene stage. Female pachytene chromosomes are
larger, and do not overlap one another as much as
the male ones do (RASMUSSEN1976; TRAUT 1976;
KAWAMURAand NIINO 1991). Recently, KAWAMURA and NIINO (1991) and SAHARA(1998) found
an asymmetrical bivalent while observing the
+
96
N . Tanaka et al.
pachytene chromosomes of oocytes in the sex-limited
yellow blood strain. They have considered this bivalent to be the Z-W bivalent.
A sex heterochromatin body (SB), deduced to be
condensed W chromosomes (TRAUT and MosBACHER 1968; ENNIS 1976; TRAUT and MAREC
1996), has been observed in the highly polyploid
interphase nuclei of Lepidopteran females. It has
been reported that the number of SBs in a cell
nucleus of B. mori corresponds closely to that of the
W chromosomes in polyploids (ITO 1977; PAN et al.
1986, 1987). Conclusive evidence for the relationship
between the SB and the W chromosome has been
obtained with W chromosome mutant strains of
Ephestiu kuehniella. Translocation or fusion of the W
chromosome with an autosome or with the Z chromosome is accompanied by fragmentation of the SB
in polyploid cells (TRAUT and RATHJENS1973;
RATHJENS1974; TRAUTand SCHOLZ1978; TRAUT
et al. 1986; MARECand TRAUT1994). The dispersed
SBs in these cases were attributed to opposing tendencies during the polyploidization of the sticky heterochromatic segment (W chromosome) and adjacent
non-sticky euchromatic chromosome segment (autosomes or Z chromosome) (TRAUTand RATHJENS
1973; RATHJENS
1974; MARECand TRAUT1994).
We have found that the sex-limited p B silkworm
strain (TWPB) has 27 chromosome pairs including an
inordinately large chromosome pair in the pachytene
stage of an oocyte, while a normal strain has 28
bivalents. In addition, we have obtained two new
strains, sex-limited and autosomal p M-like strains
(TWPML and T5PML strains, respectively), derived
from the TWPB strain (unpublished). Here, we analyze the large chromosome by using the TWPB,
TWPML, and T5PML strains. We have observed the
female pachytene chromosomes and the SBs in the
somatic cell nuclei and have made a linkage analysis
using chromosomal marker genes. The results show
the composition and the arrangement of the large
chromosome of the TWPB strain and the chromosomal constitution of the TWPB-derived strains.
Hereditas 133 (2000)
allele of it. In the TWPB strain, the females have
black larval marking, and the males have normal
larval marking. Individuals of the N21 strain are
homozygous for both pe (pink-eyed white egg, 5-0.0)
and oc (Chinese translucent, 5-40.8). This strain was
used as a marker for the fifth chromosome (linkage
group) in the linkage analysis. Both pe and oc are
recessive alleles.
The linkage analysis was based on the circumstance
that female silkworms do not have crossing over
(TANAKAI91 3). Linkage analysis for the inordinately
large chromosome of the TWPB strain was carried
out between the W chromosome with the second
chromosome fragment and the Z (sch), second (p Y),
third ([em Z e ) , fifth (pe re oc), sixth (EKp),tenth (w2),
13th (ch), or 18th (rnln) chromosomes. When a dominant marker gene was used, the segregation in the F,
was examined. When a recessive marker gene was
used, the segregation in the BF, was examined.
Pachytene chromosome preparations from oocytes
were made by following TSUCHIDAet al. (1997).
Ovaries were excised from fifth instar larvae on the
third day and immersed in a hypotonic KCl solution
(0.5 %) for 60 to 180 minutes. Oviducts were dissected from the ovaries with forceps in the hypotonic
solution and, after washing, transferred to a 1.5 ml
microtube with the KCl solution. The oviducts were
slowly fixed with an MA solution (Carnoy’s fluid)
(methanol 3: acetic acid I) and then squashed with a
polytron homogenizer (KINEMATICA). The contents were dropped on slides in a humid box, and the
slides were left overnight. The tissues were then
stained with DAPI at room temperature for 10 min
and observed under a Zeiss Axioskop microscope
equipped with epifluorescent optics.
The sex heterochromatin body (SB) was observed
in the highly polyploid nuclei of sucking stomachs. In
this material SB is easily detected in female individuals (PAN et al. 1987). Excised sucking stomachs were
dissected on slides with forceps. The tissue was then
stained by dropping acetic orcein (3 YO),
covered with
coverslips, pressed after 10 min, and observed by
light microscopy.
MATERIALS AND METHODS
We have used the strains 5137, N21, TWPB, and the
TWPB-derived strain, i.e., the sex-limited pM-like
strain (TWPMLA, TWPML) and the autosomal p M like strain (TSPML). The TWPB strain, which is
expected to be congenic to the 5137 strain ( +”/ +”)
for the W chromosome, was constructed by backcrossing more than 15 times a female of the sex-limited p B strain from S A S A K(1955)
~
with a male of the
5137 strain (OHBAYASHIet al. 1996). + P is the
normal larval marking gene, and p B is a dominant
RESULTS
The large chromosome of the sex-limited p
silkworm strain (TWPB)
In the pachytene chromosomes from oocytes, the
5137 strain of a chromosomally normal type had 28
bivalents (Fig. la), while the TWPB strain had 27
chromosome pairs (Fig. 1b) including one inordinately large chromosome pair (see arrowhead). The
large chromosome pair was about 1.3 times longer
Hereditas 133 (2000)
than the second longest bivalent, and that shape was
symmetrical and smooth. Since the TWPB strain used
here was constructed by backcrossing a sex-limited p
silkworm strain with a male of the J137 strain (n =
28), one would expect that the large chromosome of
the TWPB strain would be formed by an attachment
between the W chromosome with the second chromosome fragment (p” fragment) and another
chromosome.
We prepared sucking stomachs of the 5137 and the
TWPB strains, and inspected in detail for the presence and shape of SBs in their highly polyploid
nuclei. In the 5137 strain, a single SB was regularly
observed in each nucleus of the female moths (Fig.
2a), while no SB was detected in nuclei of the male
moths (Fig. 2b). On the other hand, in the TWPB
strain, several SBs in each nucleus (four in F i g . 2 ~and
Fusion chromosome in Bombyx mori
97
at least five in Fig.2d) were observed in the female
moths, SBs were absent from the nuclei of the male
moths. The proportion of nuclei displaying dispersed
SBs in the TWPB females was clearly higher than
that in the 5137 females. The SB in the TWPB strain
was similar to that in W chromosome mutant strains
of Ephestia kuehniella. These have a fused chromosome formed with the W chromosome and another
chromosome. Evidently, the W chromosome with the
second chromosome fragment of the TWPB strain is
attached to an unknown chromosome.
To determine the composition of the large chromosome, we crossed the females of the TWPB strain
with males of strains having marker genes on various
chromosomes, and examined the segregation in BF,
and/or F, progeny. Consequently, in the BF, progeny
obtained by crossing the female of the TWPB strain
with the male of N21 strain homozygous for both pe
and oc genes on the fifth chromosome, individuals
hatched from normal pigmented eggs [ + ”‘1 were
females with black larval marking [pq except for one
individual, and individuals hatched from white eggs
[pel were males with normal larval marking [ + P I
except for two individuals (Table 1). Accordingly, it
was evident that the large chromosome of the TWPB
strain is composed of the W chromosome, the second
chromosome fragment and the fifth chromosome. In
addition, in the F, progeny obtained by crossing the
females of the TWPB strain with the males homozygous for pe (5-0.0) and oc (5-40.8), or re
(5-3 1.7), individuals for these three kinds of recessive
genes failed to appear. This means that the fifth
chromosome of the large chromosome in the TWPB
y e , and + ‘IL genes.
strain has + P e ,
+
The strain derived from the exceptional individuals
obtained for linkage analysis on the T W P B strain
( TW P M L A )
Fig. 1. Pachytene chromosomes of oocytes. a Normal
strain (J137). Twenty-eight bivalents are observed. b Sexlimited p B silkworm strain (TWPB). Twenty-seven chromosome pairs are observed. Arrowhead indicates a large
chromosome pair. Bar = 10 urn.
In the TWPB strain there arise individuals with moricaud-like (p M-like) larval marking and detachment
of the translocated second chromosome fragment
from the W chromosome, though at very low frequency (unpublished). Therefore, we expected that
the exceptional individuals (Table 1) would be like
the pM-like males hatched from [ +
which arose as
a result of detachment of the W chromosome from
the large chromosome, and p M-like females hatched
from [pel arose as a result of detachment of the fifth
chromosome from the large chromosome. As for the
two females with p M-likelarval marking that hatched
from the [pel, we wanted to confirm the expected
detachment. We investigated the filial segregation of
characteristics by crossing the above two females with
seiae males of the N21 strain (Table 21.
98
N . Tanaka et al.
Hereditas 133 (2000)
Fig. 2. Highly polyploid nuclei of sucking stomach cells. a Female nucleus of 5137 strain. Arrowhead indicates an SB. b
Male nucleus of 5137 strain. No SB is observed. c and d Female nuclei of TWPB strain. Arrowheads indicate dispersed SB
fragments. Bar = 10 pm
The results showed that all offspring from the two
females had the [pel and the [oc], and the linkage
between the second chromosome fragment and the W
chromosome was maintained, except for six females
with the [ + P I . Consequently, the above pM-like females arose as a result of the detachment of the fifth
chromosome from the large chromosome. Among the
female pachytene chromosomes derived from batch
"b" in Table 2, 27 chromosome pairs including one
large chromosome pair were observed (Fig. 3a). Furthermore, as for the SBs in each nucleus for these
females, several SBs in each nucleus were found in
the fashion of the TWPB strain (Fig. 3b). The above
results suggested that the females with the sex-limited
pM-like larval marking arose as a result of the detachment of the fifth chromosome from the large chromosome, and by the subsequent attachment with
another chromosome.
We attempted to select from the batches for sexlimited pM-like silkworms with [pel and [oc], and we
called the batches, which had a stable expression of
p M-like larval marking, the TWPMLA strain. We
made a linkage analysis for the large chromosome of
the TWPMLA strain. This analysis suggests that an
unknown chromosome attached with the W chroniosome having the second chromosome fragment is not
a Z chromosome.
Table 1. Linkage analysis of the large chromosome of
the TWPB strain for thejifth chromosome
+P'
0
s
pe
+oc
PB
+p
PB
224
0
2*
0
1*
0
* Body color shows pM-likemarking.
OL
+p
0
212
Fusion chromosome in Bombvx mori
Hereditas 133 (2000)
99
Table 2. Segregation of larval marking and sex in the
cross of the pM-likefemales in Table 1 x p e oc males
p e oc
p M like
a
b
p s F like
Q
s
19
23
0
0
43
0
o
+p
6
F
S
10
21
0
0
4
47
53
31
0
6
100
2
Results of two batches and their sum are listed.
Sex-limited p *-like silkworm strain derived from the
T W P B strain ( T W P M L )
We subjected females of the sex-limited p M-like silkworm strain (TWPML) derived from the TWPB
strain to linkage analysis between the W chromosome
with the second chromosome fragment (p *-like fragment) and the fifth chromosome (Table 3). The results of [p*-like]:[ +"] = 1:1 and [Q]:[S]
= 1:1 in
individuals hatched from both [ +]'P and [ 9 show
that the W chromosome with the second chromosome fragment is not linked with the fifth chromosome. Also, in the female pachytene chromosomes of
the TWPML strain, 28 bivalents were observed as in
the normal strain and unlike the TWPB strain (Fig.
4a). Furthermore, in the female cell nuclei from the
sucking stomach of the TWPML strain, a single SB
in each nucleus was in general observed (Fig. 4b).
These results suggest that the TWPML strain has
originated through the detachment of the fifth chromosome from the large chromosome of the TWPB
strain.
+
Autosomal p M-likesilkworm strain derived from the
T W P B strain ( T S P M L )
We subjected females of the autosomal p M-like silkworm strain (TSPML) derived from the TWPB strain
to linkage analysis between the second chromosome
fragment (p M-like fragment) and the fifth chromosome (Table 4). The results show that individuals
hatched from the [ +""I had the p'-like larval marking, except for four individuals, and those hatched
from the [pel had the p larval marking (except for
four individuals). It is apparent that the second chromosome fragment is linked to the fifth chromosome.
This means that in the large chromosome of the
TWPB strain, the second chromosome fragment is
attached to the fifth chromosome. Also, in the female
pachytene chromosome of the T5PML strain, 28
bivalents were observed in the fashion of the normal
strain (Fig. 5a). In the sucking stomach cells of the
+
Fig. 3. TWPMLA strain. a Pachytene chromosomes of an
oocyte. Twenty seven chromosome pairs are observed. Arrowhead indicates a large chromosome pair. b Female
nucleus of a sucking stomach cell with a string of SBs
(arrowhead). Bar = 10 pm.
female moths derived from the individuals with
pM-like larval marking of the TSPML strain, a single
SB in each nucleus was in general detected (Fig.
5b). In the nuclei of the male moths derived from
the individuals with pM-like larval marking of the
Table 3. Linkage analysis of the p'-like gene in the
sex-limited pM-like strain ( T W P M L ) derived from the
T W P B strain for the fifth chromosome
p M-like
?
s
195
0
+p
pM-like
0
210
190
0
+"
0
166
100
N . Tanaka et al.
Hereditas 133 (2000)
strain is arranged in order W chromosome-second
chromosome fragment-fifth chromosome. Furthermore, this fifth chromosome contains the loci from
P" (0.0) to
OC (40.8) at least. Also, since there is
little difference in the growth rate between females
with large chromosomes and males without large
chromosomes, it is expected that the about complete
fifth chromosome of the large chromosome is attached to the second chromosome fragment. Accordingly, we conclude that the large chromosome of the
TWPB strain has originated from a reciprocal
translocation with break points close to telomeres in
both the second chromosome fragment linked with
the W chromosome and the fifth chromosome. The
chromosome constitutions of the chromosome mutant strains are illustrated as pachytene pairing
configurations in female meiosis (Fig. 6). We made a
crossing over experiment using the TSPML strain for
the mapping of the pM-like gene (data not shown). It
seems likely that the fifth chromosome of the large
chromosome in the TWPB strain is attached to the
second chromosome fragment close to the terminal
site in + O" direction.
Several cases of spontaneous chromosome fusion
have been reported in B. mori. Examples include
fusions between the sixth and 14th chromosomes
(ITIKAWA1952; CHIKUSHI1959), the sixth and seventh (SAKAIDAet al. 1996), the 23rd and 25th
(DOIRA et al. 1987; BANNOet al. 1993), the fifth
(lacking of the +p' locus) and the normal W (ONIMARU and TAZIMA1983), so far. Because of preparation difficulties, no cases of chromosome fusions have
been observed at the pachytene stage. The images of
fused chromosomes shown here are the first instances
at the female pachytene stage of B. mori.
Body colors of mutant strains derived from the
TWPB strain (TWPMLA, TWPML, and TSPML)
changed from the p larval marking into the p M-like
larval marking, and the origin of three strains was
accompanied by partial detachment of the fused
chromosome of the TWPB strain (see the Fig. 6).
Though all such detached individuals do not display
body coior variations, it is expected that the body
color change from the p B larval marking into the
pM-like larval marking shows a sign of detachment at
the fused chromosome. We shall give a detailed evaluation of multiple expression of the larval marking
due to the p locus of the translocated second chromosome fragment elsewhere.
ITIKAWA(1952) irradiated individuals having attached sixth and 14th chromosomes and obtained
3.3 % detached individuals. This detachment was
confirmed through genetic analysis. The primary
spermatocytes of these individuals had either 26 or 27
chromosome pairs. ITIKAWA(1952) inferred that this
+
Fig. 4. TWPML strain. a Pachytene bivalents of an oocyte.
Twenty-eight bivalents are observed. b Female nucleus of a
sucking stomach cell with a normal SB (arrowhead). Bar =
10 ym.
TSPML strain, SB was not detected (Fig. 5c). These
results suggest that the TSPML strain was obtained
by the detachment of the W chromosome from the
large chromosome of the TWPB strain.
DISCUSSION
The results on the TWPB, TWPML, and T5PML
strains show that the large chromosome of the TWPB
Table 4. Linkage analysis of the pM-like gene in the
autosovial p M-like strain (TSPML) derived from the
TWPB strain for the fifth chromosome
p M-like
5
8
I53
168
+p
p M-like
2
2
2
2
+"
144
136
+
Fusion chromosome in Bombyx mori
Hereditas 133 (2000)
101
Fig. 5. TSPML strain. a Pachytene bivalents of an oocyte. Twenty-eight bivalents are observed. b Female nucleus of a
sucking stomach cell with a normal SB (arrowhead). c Male nucleus of a sucking stomach cell without an SB. Bar = 10 pm.
W
-W
A
z
A
W
(In +" +'" v
+pB
Fig. 6. Female pachytene configuration of normal and mutant strains in the
present study. (a) Normal strain (5137). (b) W chromosome-pB fragment-fifth
chromosome fusion strain (TWPB). (c) W chromosome-p ""-like fragment-autosome fusion strain (TWPMLA). (d) W chromosome-p M-like fragment translocation strain (TWPML). (e) fifth chromosome-p M-likefragment translocation strain
(TSPML). W: W chromosome, Z: Z chromosome, V: fifth chromosome, A:
Autosome, (11): second chromosome fragment, *: The arrangement of the three
chromosomes (components) was assumed with n o confirmation.
102
N . Tunaka et ul.
was due to the reattachment of the detached chromosome(s) for other chromosomes. Our cytogenetic
analysis shows that the TWPMLA strain has arisen
through a detachment of the fifth chromosome from
the fused chromosome of the TWPB strain and the W
chromosome has been subsequently attached to an
unknown autosome. This is analogous to the observation of ITIKAWA (1952). We have found an instance of breakage and subsequent reattachment for
the fusion chromosome of the TWPB strain
(TANAKAet al. 2000).
In a W chromosome-autosome fusion strain of E.
kuehniellu, TRAUT(1986) reported that an attached
autosome was replaced by a different, non-homologous one. In conclusion, once a fusion chromosome is formed in Lepidoptera, and even if this
fusion chromosome is detached, it becomes easy to
form one (or two) fusion chromosome(s) involving
the two detached chromosomes. One might, at least
hypothetically, produce a series of sex-limited strains
involving a fusion of W with another chromosome.
Such sex-limited strains with probably complementary chromosomal constitutions are expected to have
rather few individuals with growth abnormalities
such as retarded growth of females in comparison to
sex-limited strains that have arisen through translocation of the chromosomal fragment on the W chromosome. Strains of this kind might be useful for the
improvement of commercial silkworms.
The TWPB strain used here originated from the
sex-limited p B silkworm found by SASAKI(1955). An
analysis of the W chromosomes in different sex-limited p B silkworm strains derived from the one described by SASAKI (1955) would indicate how the
fusion chromosome of the TWPB strain had
originated.
REFERENCES
Banno Y, Kawaguchi Y and Doira H, (1993). Cytogenetical analysis of chromosomal aberration due to translocation between the 23rd and 25th linkage groups in the
silkworm, Bombyx mori. Hereditas 118: 259-263.
Chikushi H, (1959). Contributions to genetics of Bombyx,
with special reference to mechanisms of manifestation of
characteristics. I. Sci. Bull. Fac. Agri. Kyusyu Univ. 17:
171-187. (In Japanese with English summary).
Doira H, Nakayama M, Banno Y and Kawaguchi Y,
(1987). Genetical analysis of the Naked-t mutant in
Bombyx mori. J. Seric. Sci. Jpn. 56: 220-225. (In
Japanese with English summary).
Ennis TJ, (1976). Sex chromatin and chromosome numbers
in Lepidoptera. Can. J. Genet. Cytol. 18: 119-130.
Fujii H, Banno Y, Doira H, Kihara H and Kawaguchi Y,
(1998). Genetical Stocks and Mutations of Bombyx
mori: Important Genetic Resources (ed.: H FUJII).
Silkworm Genetic Division, Institute of Genetic Resources, Faculty of Agriculture, Kyusyu University,
Fukuoka, Japan, p. 1-54.
Hereditas 133 (2000)
Hasimoto H, (1933). Genetical studies on the tetraploid
female in the silkworm. Bull. Imp. Seric. Exp. Sta. Jap.
8: 359-381. (In Japanese with English summary).
Hasimoto H, (1948). Sex-limited zebra, an X-ray mutation
in the silkworm. J. Seric. Sci. Jpn. 16: 62-64. (In
Japanese).
Itikawa N, (1952). Genetical and embryological studies on
the E-multipule allele in the silkworm, Bombyx mori.
Bull. Seric. Exp. Sta. Jap. 14: 23-91. (In Japanese with
English summary).
Ito S, (1977). Cytogenetical studies on the chromosomes of
silkgland cells of the silkworm with special reference to
the structure and behavior of the sex chromosomes. Jpn.
J. Genet. 52: 327-340.
Kawaguchi E, (1934). Chromosome behaviour in tetraploid
female of the silkworm. J. Seric. Sci. Jpn. 5: 73-79. (In
Japanese).
Kawamura N and Niino T, (1991). Identification of the
Z-W bivalent in the silkworm, Bombyx mori. Genetica
83: 121-123.
Kimura K, Harada C and Aoki H, (1971). Studies on the
W translocation of yellow blood gene in the silkworm
(Bombyx mori). Jpn. J. Breed. 21: 199-203. (In
Japanese with English summary).
Marec F and Traut W, (1994). Sex chromosome pairing
and sex chromatin bodies in W-Z translocation strains
of Ephestia kuehniella (Lepidoptera). Genome 37: 426435.
Ohbayashi F, Abe H, Simada T, Kawai S, Yokoyama T,
Oshiki T and Kobayashi M, (1996). A common random
amplified polymorphic DNA in the silkworm, Bombyx
mori is shared by W chromosomes onto which the
normal marking, Sable and Black genes are translocated
respectively. J. Seric. Sci. Jpn. 65: 395-398. (In
Japanese).
Onimaru K and Tazima Y, (1983). On a W.V translocation
strain of the silkworm with special reference to its
chromosome constitution, pairing and segregation. J.
Seric. Sci. Jpn. 52: 126-132. (In Japanese with English
summary).
Pan Q Z , Sugai E and Oshiki T, (1986). Sex-chromatin in
cell nuclei of abdominal leg of the silkworm, Bombyx
mori. J. Seric. Sci. Jpn. 55: 483-487. (In Japanese with
English summary).
Pan QZ, Sugai E and Oshiki T, (1987). Sex-chromatin
observation in cell nuclei of tissues of the silkworm
moth, Bombyx mori. J. Seric. Sci. Jpn. 56: 436-439. (In
Japanese with English summary).
Rasmussen SW, (1976). The meiotic prophase in Bombyx
mori females Analyzed by three-dimensional reconstructions of synaptonemal complexes. Chromosoma 54:
245-293.
Rathjens B, (1974). Zur Funktion des W-Chromatins bei
Ephestia kuehniella (Lepidoptera). Chromosoma 47:
21 -44.
Sahara K, (1998). Genetic and morphological features in
the polyploid silkworms induced by low-temperature
treatment. Memo. Fac. Agr. Hokkaido Univ. 21: 55-99.
(In Japanese with English summary).
Sakaida K, Banno Y , Nakamura T, Kawaguchi Y, Koga K
and Doira H, (1996). Aberrant meiotic behavior of
chromosomes due to reciprocal translocation in the ED"
mutant of the silkworm, Bombyx mori. Hereditas 125:
25-29.
Sasaki S, (1955). Genetical studies on the new sex-limited
black marking of silkworm larvae. Jpn. J. Genet. 30:
185. (In Japanese).
Hereditas 133 (2000)
Sasaki S, (1958). Studies on the breeding of a new sex-limited normal marking silkworm. Jap. J. Breed.8: 185. (In
Japanese).
Takasaki T, (1952). Cytogenetics. In: Genetics of Silkworm
(ed. Y Tanaka). Shokabo Press, Tokyo, p. 275-350. (In
Japanese).
Tanaka N, Yokoyama T, Irobe Y , Abe H, Ninagi 0 and
Oshiki T, (2000). Breakage and subsequent return for W
chromosome--pB fragment-fifth linkage group fusion
chromosome in the sex-limited p B silkworm strain
(TWPB), Bombyx mori. J. Seric. Sci. Jpn. 69: 327-330.
(In Japanese).
Tanaka Y, (1913). A study of Mendelian factors in the
silkworm, Bombyx mori. J. Coll. Agric. Tohoku. Imp.
Univ. 5: 115-148.
Tanaka Y , (1916). Genetic studies in the silkworm. J. Coll.
Agric. Sapporo 6: 1-33.
Tanaka Y, (1939). Non-disjunction of the sex-chromosomes in the silkworm. Jpn. J. Genet. 15: 359-361. (In
Japanese).
Tazima Y, (1941). A simple method of sex discrimination
by means of larval markings in Bombyx mori. J. Seric.
Sci. Jpn. 12: 184-188. (In Japanese).
Tazima Y, (1942). Cytogenetical improvement of autosexing method with use of larval marking (I). J. Seric. Sci.
Jpn. 13: 81-95. (In Japanese).
Tazima Y, (1943). Cytogenetical improvement of autosexing method with use of larval marking (I). J. Seric. Sci.
Jpn. 14: 76-89. (In Japanese).
Tazima Y, (1944). Studies on chromosome aberrations in
the silkworm. 11. Translocation involving second and
W-chromosomes. Bull. Seric. Exp. Sta. Jap. 12: 109181. (In Japanese).
Fusion chromosome in Bombyx mori
103
Tazima Y, Harada C and Ohta N, (1951). On the sex
discriminating method by colouring genes of silkworm
eggs. I. Induction of translocation between the W and
the tenth chromosomes. Jap. J. Breed. 1: 47-50. (In
Japanese with English summary).
Tazima Y, Kume T and Kamioka M, (1955). A new
method of sex discrimination of silkworm larvae by a
sex-limited moricaud marking. Reports of the Silk Science Research Institute. 5: 5-24. (In Japanese with
English summary).
Traut W, (1976). Pachytene mapping in the female silkworm, Bombyx mori L. (Lepidoptera). Chromosoma 58:
275-284.
Traut W, (1986). A genetic linkage study of W-chromosome-autosome fusions, breakage, and kinetic organization of chromosomes in Ephestia (Lepidoptera).
Genetica 69: 69-79.
Traut W and Marec F, (1996). Sex chromatin in Lepidoptera. Quart. Rev. Biol. 71: 239-256.
Traut W and Mosbacher GC, (1968). Geschlechtschromatin bei Lepidopteren. Chromosoma 25: 343-356.
Traut W and Rathjens B, (1973). Das W-Chromosom von
Ephestia kuehniella (Lepidoptera) und die Ableitung des
Geschlechtschromatins. Chromosoma 41: 437-446.
Traut W and Scholz D, (1978). Structure, replication and
transcriptional activity of the sex-specific heterochromatin in a moth. Exp. Cell Res. 113: 47-53.
Traut W, Weith A and Traut G, (1986). Structural mutants
of the W chromosome in Ephestia (Insecta, Lepidoptera). Genetica 70: 69-79.
Tsuchida K, Banno Y, Hashido K, (1997). Methods for
fluorescence in situ hybridization using oocyte and spermatocyte chromosomes of Bombyx mori. J. Seric. Sci.
Jpn. 66: 233-241. (In Japanese with English summary).