The Cultivation Of Fin Tissue And The Molecular And Cytogenetic Study Loaches Misgurnus Anguillicaudatus With Different Ploidy Status

In order to investigate the chromosome formation of loaches of different ploidy, andestablish a method of preparing chromosome using living body, in this research, natural diploid,hybridized triploid(4n♀×2n♂、2n♀×4n♂), and natural tetraploid were selected as subjects.Fin was sampled for tissue culture, and then used to prepare chromosome. Chromosome numberand karyotype of different ploidy was analyzed, and then chromosome banding was studiedusing Ag-NORs and CMA3/DA/DAPI.1. A method of preparing chromosome using tissue culture of fin cells from loaches ofdifferent ploidy10%iodine was best for sterilizing and had little influence for cell growth, and the besteffect time was15min, as cells emigrated rapidly. Chromosome number mode of all three ploidywas highest while colchicine was added3h before terminating to a final concentration of1.5μg/ml, respectively93.33%,90%, and70%. And a large number of clear methaphase spreadof moderate length, good distribution, and complete number could be acquired when treated for40min under25℃.2. Chromosome banding of loaches of different ploidy(1). Ag-NORs: number of Ag-NORs of natural diploid, hybridized triploid(4n♀×2n♂、2n♀×4n♂) and natural tetraploid was respectively2,3and4pairs.(2). CMA3/DA/DAPI: positive CMA3signals of natural diploid, hybridized triploid(4n♀×2n♂、2n♀×4n♂) and natural tetraploid was respectively2,3and4pairs.(3). Signals of Ag-NORs and CMA3/DA/DAPI of all ploidy were on the short arm terminalof the first pair of metacentric chromosome(M1), which is also the nuclear organizedregion(NOR).In sum, culturing cells of fin tissue of loaches of different ploidy could be used to preparechromosome, and chromosome number, karyotype and chromosome banding was not significantwith results using body cells. Therefore, method created in this research could be applied intocytogenetics and molecular genetics of loach researches.