O2018 Thc Japon Mcndcl Socicly C)tolosia A}Q)t 193 199

Nucleolar Organizer Regions Polymorphism and Karyological Analysis of , majusculus (Siluriformes, ) in Thailand

Weerayuth Supiwong 1*, Nuntaporn Getlakha2, Somkid Chaiphech3, Krit Pinthong4, Sumalee Phimphan5 and Alongklod Tanomtong 5

rF$ulty ofApplicd scicuc. and Enginecring, Khon Kicrr tJnive6ity, Nons Khai(nmpus, Muans, Nong Khai.13000, Thailand rD€partment ofBiology, faculty ofS(ience and Technology, Mubar Chombueng Rajabhat Uriversity. ChombuenB. Ratchaburi 70150.'l'hniland rDepartmonr ofAnimal Sui(nce. Faculw ofAgricultue. RajamanEala Univelsiry ol Tccho.r,osy Srivijaya, Nrkom Sri Tharnmdral, Saiyai Cumpus. Thtlnlsong, Naklon Si Thm naral 80110, Thliland rDepa(me,rt ofl-und.mental Scienoc, Faculty ofSuience and'ltchnok,8y, Surntdra Raj$hat tJnivErsify. Muang, Surin 32000, Thiiland 'Toxic S ub\lances in Liveslock and Aq ualic An imnls Rescarch Group. D(panment o l Biology. Foculty ofSciencd, Khon Kacn tlnivcrsity, Muang, Knon Kacn 40002, Thailand

Receiwd De.entber 21, 2017; arcetued Febtuar.t 20, 2Ul8

Summary Nuclcolar organizcr regions (NORS) and karyotypc in thc blaok lanu(J, Bagrichlhts tnaj $culus fiom Nakhon Phranom and Sing Buri provinccs. Thailand, wcrc invcstigatcd. Thc nlitotic chromosomc prcpara- tions wcre directly obtained from kidney tissues of eight male and eight fcmale spccinrens. Conventional and Ag-NOR staining tcchniques wcrc applied to the chromosomcs. The chromosomes numbcr of B. ntojlsc lus \Nas 2n:50 and the fundnnrcntal numbel (NF) ofall specimens was 96. The klryotypc consists ofsix large metacen- tric, six large submctaccntric, two largc acrocentric, eight medium mctaccntric. l2 mcdium submetacentic. two Nedium acrocentric. six sm0ll nretacentric, four sruall subllretaceDtric und four small teloceotric chtromosoDes. Scx chromosome was not obscrvcd. Morcovcr. thc intcrstitial NORs wclc clearly obscrved at the long arn1 ofthc metaccntric chromosome pair l. Th('rcsult levcaled that a hcleromorphic NOR typc was appearcd in onc malc and onc ttmale and other fish have homozygous NOR in the pair I chromosomcs.

Key rlords Ragrichth.v-s majusdrh.r, Karyotype, NOR variation. Polymorphisrn, C-hromosomc nrrnber

Freshwater are especially important as thcy of and interrelationships within fish families providc a sourcc of high quality protcin as wcll as food (Campiranont 2003, Taoomlong 20ll). Thc studics of source lbr peoplc who livc near a river basin. Fishes of the karyotypes help to invesligate thc aquatic structure the Bagridae tbmily belong to the order Siluriformes and of the species population in each habitat, so it can de- are of great importance as fbod lish. While. sorte spe- termine what species are related to each other in an ac- cies are kept as aquarium 6slres, nrany bagrids have re- curate manncr. This may hetp to facilitate the hybridiza- cently bccomc a boorning iu aquaculturc (Nclson 2006). tion bctv,,ccn them in thc f'uturc tbr strain improvcrnclt Therc arc six gcncra and approximatcly 25 spccics in thc (Sofy e/ a/. 2008), brccding practiccs of organisms by Bagridae family in Thailand (Rainboth 1996, Vidthaya- using chromosome set managcm€nt such as polyploidy non 2005, Ferraris 2007). B. maju.tculus is onc spccics inducing in catfishcs, Clarias bdtrdchu,t (Na-Nakorn ol thc lumily considcring as thc Tltai ecunolniu spccics et ol. 1980\ and rainbow tto\l loncorh.vnchu-t ,[,kis.t), rused for foods and in an aquarium (Vidthayanon 2005). brood stock selection of giant (MengamPan el 41. Thc karyotypc is thc chromosourc con'tplcnrcnt of an 2004). Cytogcnctic knowledgc can also providc increas- individual or related group of intlividuals, as defined by ing useful data for future studies on chromosome evolu- chromosome sizc, morphology and number. Though for tion. The rcports of chromosome evolution in thc family all somatic cclls ofall individuats ofspccics, thc numbcr Channidac (snackhcnd 6sh) indicatcd that ccntric f'usion, of chromosonrcs is used as an indicalor of classification pcriccntric inversion and polyploidy ars the processes which have important roles on chromosome rearrange- rnent in the snackhead fish's phylogeny (Supiwong el a/. * Corrcsponding uulhor, e-mai! : supiwong(rghot,nail.eom DOI: 10.15o8/cylologia.83. 193 2009, Tanomtong e/ d/. 2014). I9.l Cytologia 83(2)

arc oftcn ttscd Chromosqmc number and karyotypc A N as cssential data for the of those with troubles on morphological classitication aspect (Giessmann 2002). tn fishes, chromosome banding data was used to support for classificatio[ in lhc CyPrinidac (Amemiya and Gold 1988, Gold and Li 1994, Boron 2001, Bcllafrontc e/ d/. 2005). Up to the present. 46 species of the Bagridae have bccn cytogcnctically studicd. The chromosome numbcr ranging from 2n=44 in Coreobogrus brevicorpus (Kint et ol. 1982) to 2rr=80 in Butusio havnolleri (Magtoon and Donsakul 2009) have been reported. However. the predominant diploid numbers represent 2r:52 and 2r=56 (Arai 20ll ). In the present study, chromosolne of thc B. najus.ulus ftorn Thailand was firstly investigatcd using conventional staining and Ag-NOR banding tcch- niques.

Matcrials an(l mcthods

The specimcns of B. najusutlus wcrc collected from 0 the rivcr, Nakhon Phranom province, northeasl = Sam pling Sites of Thailand (fivc males and five fcmalcs) and thc Chao I Phraya Rivcr, Sing Buri province, ccntral part o[ Thai- land (three malcs and $ree f'emales). Thc sampling sitr.'s o B and the characteristics of B. mojrccuhr.r are shown in l'... . Fig. l. The fishes were transt'erred to laboratory aquaria and were kept under standard conditions for seven days prior to the cxpcriments. Proccdurc of chromosornc prcparation was following modified protocol of Supi- wong et al. (2009). The 0.05% colchicine (l mL to l00g 0 body weight) was injected to abdominal cavity and lcft for 45 min to I h. Chromosomes were prcpared from kid- ney cells by squash technique. Kidney tissues were cut Fig. l. Thailar mnp showirg lhe s.rnpling siles (A) rnd the gen- into small picccs, then mixcd with hypotonic solution cral charactoristic of8- ,r(rrlrrcllrr.r (B). (0.075 M KCI) and incubatect for 30rnin. Cells were fixed in fresh cool fixative (3 absolute methanol: I glacial ace- et aL1989, Lakra and Rishi 1991, Khuda-Bukhsh el o/. tic acid). The propared chromosomcs werq staincd with 1995, Donsakul 2001, Das and Khuda-BLrkhsh 2007a, I0% Giemsa tbr 30min, or 50% AgNOi and.2Yo gcla- Supiwong et a1.2013a,2014a, b). The NF of B. majrrs- tin lbr Ag-NOR staining technique (Howell and Black culus was 96 in both males and t'emales. Although our 1980). The metaphase figures were analyzed according result presented the same chromosome number as in to the chromosome classification following by Turpin previous studies, the NF is different (Donsakul 2002, and Lcjcune (l9651and Chaiyasut (1989). Magtoon and Arai 1988). Donsakul (2002) rcported that B. majusculus showcs NF=92 whercas Magtoon and Results and rliscnssion Arai ( 1988) reporled NF=94. These differences may be duc to different critcria uscd tbr the chromosome clas- Chromosome number, fundantetttal nunber and kar.yo- sificalion and/or inter-populalional variation in this spc- ,,pe o/8. majusculus cies. The NF in the family Bagridae varies tiom 72 to Cytogenctic studics havc bccn pclfornrcd on sixtccn 128 (Arai 201 l). Comparisons among the gcncra in thc specimens of B. naiusculus from Thailand. The results Bagridae revealed that the Hemibagtus showes higher rcvcaled thal thc diploid chromosontc numbcr was NF variation than others. lt varies from 72 to I14. Ghi- 2n=50, in both males and fcmalcs (Figs. 2, 3). This is gliotti et a/. (2007) assumcd that spccics with a largcr accordancc with the prcvious studics of Magtoon and NF are more advanced in evolutionary terms. Such Arai (1988) and Donsakul (2002). However. the 2a is changes in NF appear lo be related to the occurrence of diflerent from other species of the Bagridae (Barat and pericentric inversions, which are among the most com- Khurla-Bukhsh 1986, Sharma and Tripathi 1986. Yu mon nrutlificalions cuntributing to karyotypic rearrangc- 2018 NOR Polymorphism.nd Krryologicrl Anitlysis uf Bugtk hthr! ttfur{ulux in Thailand 195

I{.Ir' rr rr rr rr r. ll lI rt rr rr rt .r .r ml r 2 ! a 5 6 't7 t lr3as51a tt0 910 la II TT rl ll aa la tI II r,l t, lll4 r7 lt "t 1l l2 t3 td l5 t6 l? t8 lt t9 l0 2t t9 70 2l r lrr al 23 77 2-1

24 25 24 15 B

FiB. 2. Karyotypes of male (A) and fcmale (B) of B. ,nai$culus (2n=50) by convcntional straining tcchnique. Arrows indicalc seoondary conslriction. Scale bars=5tlm-

Tsble l. Mean length (,!m) ofshort chromosome arm (Ls), mean length oflong chrumosornc ann (Ll). total lcngth olchromosome anns (LT). rclarivc lcnglh (RL). ccnlromcric ilxlcx (Cl) and slandard dcvi.tion (S.D.) of RL and Cl from mctaphlsc chromosomcs of 20 cclls ;n B. nuj6crhts (2n=so).

Chromosomc pail Ls LI L'I RLIS D CllS.l) Type Sizc

47.59 55.45 103.04 0.029410.0022 0.519+0.027 L 2 19.39 5r.05 90.44 0.0157r0.002.1 0.566i0.0-'t2 L 3 34.80 44.59 79.18 0.022510.0019 0.560i0.030 L 4 31.0-'i 19.65 7t.68 0.010510.0009 0.553r0.010 M 5 30.14 67.41 0.0112i0.0011 0.554t0.02t) M 6 29.4t 34.85 64.26 0.0183 i0.000(, 0.544r0.028 M 1 27 _96 33.21 6!.t7 0.0174i0.000it 0.542L0.1)26 M I 24.90 3t.37 56.28 0.0161 10.0012 0.559r0.029 m s 9 23.15 29.10 52.25 0.0150r0.0012 0.557+0.024 m s t0 20.64 27.X3 4ti.46 0.0138r0.0011 0.57 5 !,0.022 s ll 36.98 76.48 I13.46 0.012110.0017 0.673r0.058 L t2 13.98 62.14 96.12 0.017510.0020 0.647 rO.029 sm L ll 27.O9 5?.01t 84.t1 0.023910.0{rll o.619rO.022 L t4 26.92 50.79 17.7 t 0.022110.00ll 0.654r0.028 M l5 26.t2 47.13 71.85 0.0210j0.0009 0.641!0.021) M t6 24.O9 46.49 70.59 0.0201 i0.0007 0.660r0.0:i5 M t7 24.4t 4 t.84 66.25 0.0,89a0.0011 0.62610.030 f,4 t8 22.66 40.96 63.62 0.0ltl I r0.(xm6 0-65110.0:E M t9 21.94 18.82 6t).76 0.017i10.0007 0.61810.030 M 20 20.6t 16.09 56.70 0.016010.0012 0.637r0.032 s 2l 17.02 35.00 52.O2 0.01.1710.0015 0.674r0.042 S 22 21.94 57.59 80.51 0.0130r0.0015 0.714r0.026 L 23 t.62 49.21 67.1t9 0.0193+0.0012 0.72Iir0.031 a M 24 0.00 55.t'0 55.60 0.0159r0.001t 1.000i0.000 t s

0.00 42.a3 42.83 0.0122+0.0012 L000*0.000 1 s

'NOR-bcaring chromosomo, m=mclnccntric. snr:submclaccnlric, a - icroccnl ric, I - I ckrccnl ric, L:largc. M=mcdium, S=snall ment in fishes (Galetti et ol. 2000, Wang et al. 2010) and is similar to other species in the Bagridae of Thailand other vertcbratcs (King 1993). (Donsakul 2001, Magtoon and Donsakul 2009, Supi- Th() karyotypc ol B. mdiusculus composed of six wong et al.20l3a, b, 2014a, b). However. somc bagrid large metacentric, six large submetacentric, two large in lndia, including M. gulio and M. tengara acrocentric, eight medium metacentric, l2 medium have differentiated sex chromosome system as XX/XY submetacentric. two modium acrocantric, six small antl ZZ/ZW, respectivcly (Rishi 1973, Rishi and Rishi metacentric. four small submetacentric and four small 198 t, Choudhury e, al. 1993, Arai 20ll\. telocentric chromosomes or 20m+22sm+4a+4t (Tablc l). The present result is disagreement from Don- Ch ronosome marker.r o/8. majusculus sakul (2002) and Magtoon and Arai (1988) report€d The Ag-NOR staining technique identifies the NOR as 2r=100=20m+14sm+8a+8t and l6m+26sm+2a+6t, which is thc rcpresentativc location of active l8S and respectively- These differenccs may be causcs from 28S ribosomal RNA gencs (Jordan 1987). Atlcr Ag- the intraspecific variation, since they represent dif- NOR banding the chromosome in B. nwju"'culus re- f'er€nt populations. All investigated specimens had no vealed the positive Ag-NOR marks on a single pair of distinguishable scx chroorosonle. This characteristic the largest chromosone pair l. The Ag-NOR was local- t96 Cytologia 8l(2)

'Iiblc 2. Numbcr of mctaphase cellswhich showing honlomorphic or lt lr rr a, . at heleromorphic Ag-NORs on the chromosome pair I of B. m 5 6l I ajus.utts. lx la al234 g l0 observed Cells with Cells wilh Specimen Sex cells (No.) Ag-NOR (No.) A8-NOR (No.) sm ll tl t-1 ll l5 I6 t1 t I Male t0 t0 0 l9 20 ll 1 Malc l2 t1 0 l Male l5 r5 0 I la. I Mnle 20 20 0 2-1 5 Malc 20 0 20 l6 l6 0 r l.- A 6 Mnls 24 25 1 Male l5 I5 0 8 Male l8 l8 0 l5 l5 0 l0 Female l5 l5 0 nl llt Iltlrtrr lr lc 2 345 6llt ll Fernale 10 l0 0 t.a l: Femtll€ 16 t6 0 9 l0 l3 Fernale l5 l5 0 t4 Female :0 20 0 stD l,t ,a t5 |'cmalc 2t 0 2l t2 l3 IJ 15 l6 l1 llt lt !-cmalc t7 t1 0 l9 t0 2l Torsl= l6 255 211 . "lr I .t t B gous chromosomc of pair I was also cxhibitcd in onc ll 25 mrlc tnd onc l'cmalc specimens. Two spccimcns had a singlc clrromosome of thc chromosome pair I (lalb), I ra while seven females and seven males had two NOR- m tt ll rr rr rr lr lr lb 2 74567 ^ t ll al bcaring chtomosomes of the pair I with a homomor- 9 l0 pltism (lala) (Table 2, Fig. 3). Ag-NOR banding can dctcct protcins associatcd with IDNA cistrons that arc Irt activcly transcribing rRNA for nucleolus production in sml I t2 l3 t.1 t5 t6 t1 1.. lhe previous interphas€. When the NOR-bearing chro- t9 l0 ll mosomcs condcnsc and entcr prophasc. thcsc rRNA- associalcd proteins are entrapped around the NOR antl " lrr t.r it is these proteins that are still present in the metaphase chromosomc that selectively rcduce the silvcr and bc- ,lrrt C 25 cornc heavily stained (Howell 1977). whereas unstaincd Fig.3. Karyolypes ofmale (A) and lbmale (8. Cl of R. itojusculus NORs did not transcribe rRNA during the preceding (2a=50) by Ag-NOR banding technique- The chromo- intcrphiNc and thcrc are I]o or lcss rRNA-associated pro- some lal! show as homomorphic NORS (A, B) while thc tcins, hcncc no silvcr stain (Howell and Black 1979). Ac- chromosomc lalb show as heteromarphic NOR (C). Scalc bars=51,m. cordingly, the lact that Ag-NOR on one chromosome oI the pair I indicated that another one might be inactive. This intraspecific NOR pulymorphism is cumrnon cvcnt izcd at intcrstitial rcgion of thc long arm in thc largcst lbund prcviously in scvcral lishes in Moenkhqusiu sunc:- mctaccntric chromosomc pair I (Figs. 3,4). Thcsc marks toeflomenae (Forcsti el a/. 1989), Aphaniu.s fascians are in accordance rvith the secondary constriction. The (Vitturi c, al. 1995), Leporinus -frideri<:i (Galetti et a/. chromosomc pair I carrying Ag-NOR is consistent with 1995), Salmo tutto (Caslrc et al. 1996), Salvelinus al- H. nrenodanenodq (Barat and Khuda-Bukhsh 1986), M Trlrres (Rccd and Phillips 1997\, Chondrostonta lusitani- 6oc.,x/.li (Supiwong et ul.2013a), Pseudon.vslus tiame - crim (Rodrigues and Collares-Pereira 1996, Collares- srs (Supiwong et ul. 20l3b), Horabagrus btathvsotna, Pcrcira and Rib 1999). Hoplias makboritus (Born and Ho. nigricttlluris. M. cuvusius. M. vittutus, Peheobagrus Bertollo 2000), Oeddec'hilus /aberr (Rossi eI ul.2O0t)). ussuriensis, Pseutlobugrus vuchellii (Arai 20Il), M. gu- Aslyunux scobripinris (Mantovani et.l/. 2000, Marco- lio, Rita ritu (D'ds and Khrrda-Bukhsb 2007b) and Spcru- Fcrro et dl. 2001, Soza et al. 2001), A. ultipurunae la seenghola (Das and Khuda-Bukhsh 2007a). Howcvcr, (Pachcco et al 2001. Mantovani et al. 20O5), Bqycorr it is ditferent l'rom Ta<'hvsurus ./iividraco which has two .rrreri( r.r (Paintner-Marques e, a/. 2OO2\, Aparekxktn aJ- pairs of NOR-bearing ch romosomes (Arai 20 I I ). Jinis (Jorge and Filho 2004), Aphanius lAsciotus lVitluti Intraspecific NOR polymorphism between homolo- et aI.2005), Prochilodus lineatus (Gras el a|.2007), Br. 191 2018 NOR Polymorphis .lnd Krryological An lysis of BuSthhthys &jLtr.ultts in Thailand

Frurd lbr DPST Graduatc with First Placement Year 2015, De!'elopment Promotion of Science and Tech- ;/ nology Talents project (DPST), scholatship under the Post-Doctoral Training Program tionr Research Alfairs lltltt rrrr lllttllr and Graduate School, Khon Kaen University, (Crant no. 59255) and Toxic Substanccs iD Livcstock and Aquatic Animals Research Group, Khon Kaen University, Thai- land. Iltn rr u, llllllrru Rel'cr cnccs

Amemiya, C. T. and Gold, l. R. 1988. Chromosomal NORS as taxo- !r uu nornic and systcmalic characlers in North Americin cyprinid fishcs. Gcnctica 76: 8l-90. Arai. R. 201l. Fish Karyotypc: A Chcck Lisl. Springcr. Tokyo. Fig. 4. Idiograms shorving lengths trrd shrpes of chromosomes of Barat. A. and Kbuda-Brkhsh. A. R. 1986. Karyomorphometrical B. naJuscuh$ (2r-50), by convcntioMl sttlining {A) and studies ir) tlvo spccics of fishes, L(pklucephali.hthys gttutca As-NOR brnding tcchniques (B). Arrou'ind;otes Ag-NOR. (lam.: CobitidnB) and r -s/r?.r cortrld (iam.: Bagridae). PersPccl. Surlc bars=l tnr. Cylogeret. Celt Cenel.5: I l5-l18. Bellafronte. E., Margarido, V. P rnd Moreira-Filho, O.2005. (Capislaro et ul. 2008), Puntioplites ptodol)'s- iheringii Cytotaxonomy ofPdro./rn nasus and Purolo brlrlrsr.! (Pisccs, roz (Supiwong et ol. 2Ol2), Lutja us johtlii (Phimphan Characiforncs). A casc of synonynry conGmcd by cytogcnctic ct al. 2013). Pterapogon kauderni (Kasitock et al. 2017\ Jn:rl).cs. GcIcl. Mol Biul.2E: 710-716. atl ltemibagnrs r),cf,li (Supiwong et tl. 201'7). Therc- Born- G. G. and Bertollo. L. A. t-.2000. An XX/XY sex chromo- wit|r a forc, difGrcnt karyotypcs arc tbund cven in stnall and som€ syst€m iD i fish speries. Hoplia: uldhutitus, polynrorphic NOR-bcaring x chroml,some. Chronrosonre Res. isolatcd populations of thcsc specics (Cdpistano e/ o/. 8: lll-118. 2008). Thc use of NORs in explaining kinships depends Boron, A. 2001. Comparative chromosomal sludies on two minnow on a large extent on the unilbrmity of this characteristic 6sh, I'ro.rr.nr (Linnaeus. l75a) Eupdllosellu l,ro-rr',(r ^nl and on the tlegree of variety within a taxoo (Yiiksel and /,erzrrrrr (Pat1as, l8l4); an associalcd cytogcnctic-laxonomic Cafl'aroElu 2008). lonsideralion. Oenelica lll: 387 i95. Campiranont, A. ?003. CytoSenetics. 2fld ed . Depcrlment ofGenet- All spccimcns investigatcd in thc prcscnt study havc ics, F:€ulty ofScicncc, Kasctsart Univcrsity, BanSkok. (n/ Irar) largest mcta- thc interstitial Ag-NOR on long arm of the Capistano, T. C., Porlela Castro, A. L. B. and Julio-Junior, H. F. 2008. centric chromosome pair l. The interstitial Ag-NOR in Chronresome divergence and NOR polyrnorphism in Bf,coa this species is similar to P. sianensis populations from unrctit t aff. ir.,rrgii (Teleoslei, Chara€idae) in th€ hydro- the Songkham and Chao Phraya Basins (Supiwong er a/. graphic sysrems of the Paranipanem. and lva; river, Parand, Brazil. Gcnct. Mol. Biol .31 2031111. 2013b) and rI r):cklr (Supiwong el al.2Ol7), but there is Castro, J., Viiias, A.. Sincbez, L. ard Marlincz, P. 1996. CharacteF rcsult the River population that they ditfcrcot from Chi izatioo of ar alypical NoR site polymorphism in brown trout pose telomeric Ag-NOR (Supiwong et a/. 2013b). The (Jrn1,n,,.l/,ll,) with Ag- a CM^.-staining, and ffuoresce t ir. important chromosome marker of the B. majusculus is rirr hybridizrition. Cytogenel. Cell Cenet. 75: 234-239. thc asymmetrical karyotypc that all four typcs of chro- Chaiyasut, K. 1989. Cytogenclics nnd Cytolaxonorny of the Family mosomes (metacentric, submetacentric. acroccntric, and Zephyranlhes. Departmenl of Botany, Faculty ol Scicnce, Chul' aloDgkorn Univcrsily, Bangkok. (i" ftai) telocentric chromosomes) were presented. An idiogram Choudhury, R. C., Prasad, R. and Das, C. C. 1991. Chromosomcs of displays a continuous length gradation of chrornosomes. four I0diau marine catfishes (Bagridae. Ariidae: Siluriformes) The largest and snallesl chromosomes show size tliffer- with $ hetcromorphic pair irl mnle M),srzs -srln. C.rryologia 46: cnce approximately 2.5 tblds. Thc markcr chromosomcs 231 243. poly of B. majusctlus arc thc chronosomc pair I which is Collares-Pereira, M. J. aad RAb, P 1999. NOR orphism in the lberian spccies L'hondnsto,ttt lusrl.rri.rr, (Pisces: Cyprinidae) the Iargest metacentric chromosorne and chromqsome re-examination by flSH. Genetic.t 105: 301-103. pair 25 which is thc smallest tcloccntric chromosomc. Das, J. K. .rnd Khudr-Bukhsh, A. R. 2007a. Gc-rich hel€rochromatin The data of the chrormrsomal chccks on mitotic mola- in silver staincd nuclcolar orsanizer resjons (NORS) fluoresccs phase cells of B. majusculus are shown in thble l. The with chromomycio Ar (CMAr) sta,ning ifl rhree species oftcleos- idiograms from conventional staining and NOR banding tean fishes (Pisces). Indian J. Exp. Biol.45:413 418. Dns, J. K. and Khuda-tsukhsh, A. R. 2007b. Preponderance ol GC- techniques are prisented in Fig.4. The karyotype for- rich sitcs in silvcr slaincd n clcolus organizing rc8ions of Ril, nujus<:ulus is follows: 2,7:2,r=50=Lll+ mula for B. as ,'ra (Hamiltor) and M$/ur grl/io (fiamillon) (Bagridae. Pisces), Ll'+L;+N4i'+Ml+Ml+Sr+Sl''+S; ns rcvcalcd by chromomycin Ar-slairing techniqu€ .rnd sc.rnning el€ctron microscopic studiss. Genct. Mol. Res. 6: 281 29i. Acknowledgements Donsakul, I 2001. 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