I

Asiatic Herpetological Research

December 1993

Vol. 5, pp. 45-50

Karyotype, C-Band and Ag-Nors Study of Three Stink Frogs

Gang Wei ning Xu dejun Li Guanfu wu 2 and xiquan Song3
11,1,,'Department of Biology, Zunyi Medical College, 563003 Guizhou, China
2Chengdu Institute of Biology, Academia Sinica, 610015 Sichuan, China*Department of Biology, Zunyi Teachers College, 563002 Guizhou. China

Abstract.-The karyotypes.C-bands and Ag-NORs of Rana kuangwuensis, R.andersonii and R.margaratae

were analyzed. Intra- and interspecific chromosome variations, including centromeric type and C-banding
patterns, were detected. It was assumed that the Guizhou Plateau was the distributional center of the original place of the group.

Key Words: Amphibia, Ranidae, Rana kuangwuensis, Rana andersonii, Rana margaratae, China, karyotype, C-band, Ag-NORs.

U«M«K"*» lYltfXXftlAAAA•t}f!Mtt(FIG. 1.

a:Karyotype of Rana andersonii.

,t, « tt|,|b:showing C-bands.

Introduction

c:'*M

showing Ag-NORs.

NORs of R. margaratae from Emei MounWangtain, Sichuan have been analyzed byThe group of stink frogs which have aspecial stink smell from the skin consists ofnine species, i.e. Rana andersonii, R. anlungensis, R. grahami, R. kuangwuensis,
R. lungshengensis, R. margaratae, R.
schmackeri, R. tiannanensis and R.
wuchuanensis. They are considered to bephylogenetically close because of similarmorphological characters in adults and tadpoles. Among them, the karyotype and Cbands of R. grahami from Kunming, Yunnan has been studied by Li (1982). In addition, the karyotype, C-bands and Ag-

1993 by Asiatic Herpetological ResearchWu(1990). In the present pa(1983) andC-bands and Agthekaryotypes,per,NORs of R. kuangwuensis, R. andersonii
and R. margaratae were analyzed.

Methods

Two females and one male R. andersonii
Qianxi, Guizhou
Province, (27°20' N,106°16' E). One female and three male R. kuangwuensis werecaptured at Nanjiang, Sichuan Province(32°30' N, 106°40' E) and one female andwerecapturedat

Asiatic Herpetological ResearchVol. 5, p. 46

11 ' 1^{((IKXilYAI!

v

f.

ifDecember 1993

" 1 "" 1" atH h }]

,|mmw
mMm^

r vtffVWM*ikRG.2.

M .1 •»a:^Karyotype of Rana kuangwuensis.

b:showing C-bands.

c:AAshowing Ag-NORs.

one male R. margaratae from Zunyi (27°40'N, 106°50' E) were captured. Karyotypes,C-bands and Ag-NORs preparations wereThe small chromosome group comprises
chromosome Nos. 6-13, with a R.L. lessthan 7%.
Nos.6, 8, 10, 12 and 13 aremade after Wei et al. (1990).

metacentric, No.7 is submetacentric in all
the three species. Nos. 9 and 1 1 are submetacentric in R. kuangwuensis and R. andersonii but metacentric in R. margaratae.
Secondary constrictions are observed in thelong arms of No. 10 of R. margaratae (onlyResults
Figures

1,2,and3

thedepictkaryotypes, C-bands and Ag-NORs of/?.
andersonii, R. kuangwuensis and R.
margaratae. For the measurment of thekaryotypes see table 1 The diploid numberof the species are all the same, 2n=26,
comprising two groups.

one homologous) and R. andersonii but notobserved in R. kuangwuensis.

.

The large chromosome group includes
chromosome Nos. 1-5, with a relative
length (R.L.) larger than 9%. With regard
to the arm ratio (A.R.), chromosome Nos.

Treatment of the chromosome of thethree species according to the C-banding
method shows that each species has acentromeric C-band on each chromosome.

For interstitial C-band, it is quite different
among the species. There is only aninterstitial C-band inlOq in R. andersonii.

and 5 are metacentric in all three species.
No. 2 is metacentric in R. margaratae andR. andersonii, but submetacentric in R.
kuangwuensis. No.3 is submetacentric in
R. margaratae and R. kuangwuensis butmetacentric in R. andersonii. No.4 is submetacentric in R. andersonii and R. mar-

And there is an interstitial C-band in 2p

garatae, but metacentric in R. kuangwuen-

kuangwuensis has some telomeric C-

sis.

bands.

1,(stained weakly), 3q and 4q (only onehomologous) in R. margaratae. But there
are much more interstitial C-bands in R.

than in the other twoR.
margaratae and R. andersonii
species.
have not any telomeric C-band. But R.

kuangwuensisAsiatic Herpetological Research

December 1993

Knuunxu",!

|^ KK]J ]i|5FIG. 3.

a:Vol. 3, p. 47

lIAXtlAXfti

iiwii it u •« ii

Hi M'* II »Karyotype of Rana margaratae.

b:showing C-bands.

Specific staining of the NORs with silver(Ag) confirms that the regions of NORs in
all the three species are the same, in thec:*iI

showing Ag-NORs.

But thebetween the NORs and the con-margaratae are similar to those of R. andersonii, except for the variant bands onchromosomes 2, 3, 4, and 10. While Cbanding pattern of R. kuangwuensis isquite different from those of the other twostrictions is quite different. The regions ofthe one pair of NORs in R. andersonii cor-species, for R. kuangwuensis has more interstitial C-bands and telomeric C-bands as

respond to the regions of the secondaryconstritions. R. margaratae has only one
NORs in the chromosome where the secondary constriction locates, and no NORs
is observed in the other homologous which
has no secondary constriction. While forR. kuangwuensis, no secondary constriction is observed in the regions of thewell.

long arms of chromosome No. 10.
relation

NORs.

And the relation between the NORs
and the secondary constrictions is quite different among the three species.

Thisstudy

also

indicatesthat

intraspecific chromosome variations exist in

R. margaratae from different distributional
observe 8 metacentric and 5areas.
submetacentric pairs in the present study,We

opposed to 12 metacentric and 1
submetacentric pairs from Emei Mountain,

asDiscussion

et al.,
1983) and 94and
submetacentricmetacentric
pairs also
from Emei Mountain (Wu 1990).

SichuanComparing the karyotypes between thethree species, we detect some interspecific

The secondary constriction iskuangwuensis. The
chromosomes consist of 8 metacentric and(Wangvariations.

not detected in R.

5 submetacentric pairs in R. kuangwuensis

and R. margaratae, while 9 metacentric and
4 submetacentric in R. andersonii.

C-banding patterns of all 13 pairs of R.

The C-banding pattern of chromosomes
1-13 of R. margaratae from Zunyi is compared with those from Emei Mountain.
There is a centromeric C-band in eachchromosome and a terminal C-band at eachchromosome terminus, and an interstitial Cband in the long arm of No. 3 from Emei

Vol. 5 p. 48

TABLE 1.

Asiatic

HerpetologicalResearch

Karyotypic data for Rana margaratae, R. andersonii, and R. kuangwuensis.

Chromosome Nos.

December

1993

December 1993

R.

anlungensis:
County)Asiatic Herpetological Research

Guizhou (Anlungsumed that the stink frog group originatedin Emei Mountain and its adjacent plateau.

R. grahami: Sichuan, Guizhou, Yunnan

R. kuangwuensis:

We are most grateful to Ms. Wei Wu for

Guizhou, Guangxi,

providing us with a pre-publication
manuscript and Ms. Xiaomong Zheng forhelp in collecting specimens andexperimental work. This research wassupported by the Guizhou Provincial
Committee of Science and Technology.

Hunan
R. margaratae:
GuizhouAcknowledgments

Sichuan (NanjiangCounty)
R. lungshengensis:

Vol. 5, p. 49

Gansu, Sichuan,

Literature Cited

R.

schmackeri:

Henan,Gansu,Sichuan, Guizhou, Hubei, Anhui, Jiangsu,FROST, D. R.

Zhejiang, Jiangxi,Hubei, Guangdong
R. tiannanensis: Yunnan, Hainan

R. wuchuanensis:

IKEBE, C, A. KEN-ICHI and K. SEI-ICHI.
Karyotype analysis of two Japanesesalamanders, Hynobius nebulosus (Schlegel) andHynobius dunni Tago, by means of C-banding.
1987.

Libo)

Zoological Science 4:621-626.

From the discription above, it could bethat the stink frog group is distributed
mainly in the south of China, and most ofthem (7 species) are found on the Guizhou
Plateau. So, the Guizhou Plateau might be
the distributional center of the group.

There were another two species of stink
Their karyotypes and C-bandingfrogs.
patterns were published. They are R. grahami and R. schmackeri. Both the species
have 10 metacentric and 3 submetacentric
pairs in their karyotypes, and both species
have one telomeric C-band, but the former
has 5 and the latter has 4 interstitial C-

Amphibian species of theThe Association of Sys-

tematics, Lawrence, Kansas.

Guizhou (Wuchuan,

found that the distributional areas of somespecies are very limited, only one or twocounties. So they are very rare and precious wildlife. And it could also be found

1985.

world. Pp. 480-520.

LI,GUOZHENG.

1985.

research method.

Science Press, Beijing Pp.l-Chromosome and its

261. (In Chinese).
LI, S., Y.

WANG, C, LI, R. WANG and G. LIU.

1982.

An investigation for the karyotypic andC-banding pattern on the two anuran Amphibia.
Acta Genetica Sinica (6):473-478. (In Chinese).
1978. Chromosome banding inConstitutive heterochromatin andSCHMID, M.

Amphibia.
nucleolus organizer regions in Ranidae,Microhylidae and Rhacophorodae. Chromosoma
68:131-148.

WANG, Z., X, WANG, and W. CHEN.

Acta Herpetologica Sinica 2(4): 1-6.

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A

1983.

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comparative(InChinese).

Among the 5 species published theirkaryotypes and C-banding patterns, R.
margaratae from Emei Mountain has mostmetacentric pairs and most telomeric Cbands. Although it does not have less interstitial C-bands, it could still be considered as the most original in the viewpoint ofcytogenetics. Considering that R. margaratae from Guizhou is more evoluted thanthat from Emei Mountain,

itmight be as-WEI,G., F.

CHEN, and N. XU.

1990.

An

investigation of the karyotypic C-banding andAg-NORs pattern on Rana chensinensis.

Hereditas (Beijing) 12(l):24-26. (In Chinese).

WEI,G.,N.

XU, andD.

LI.

1990.

An

investigation of the karyotype, C-banding, Ag-NORs

on Rana schmackeri.
patternand Reptiles,ofAmphibians
Cytogenetics
Advances in Life Sciences. 147-152. Birkhauser

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Huey, R. B.

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