多倍体泥鳅的人工诱导及其受精细胞学观察

Olyploidy induction and fertilization cytology observation in loach

对大鳞副泥鳅(PP)、二倍体泥鳅(DD)和四倍体泥鳅(TT)自交或杂交受精卵进行冷休克(冷休克条件为:受精后5min开始,利用2~3℃冰水冷休克处理30~35min),获得6种多倍体泥鳅后代(DD×PP-0,3n=74;DD×DD-0,3n=75;DD×TT-0,4n=100;TT×PP-0,5n=124;TT×DD-0,5n=125和TT×TT-0,6n=150。母本在前,父本在后)。利用流式细胞仪对不同诱导组合不同时期倍性进行检测发现:1月龄,DD×PP-0三倍体诱导成功率为87.50%,DD×DD-0三倍体诱导成功率为53.85%,DD×TT-0四倍体诱导成功率为90.91%,TT×PP-0五倍体诱导成功率为84.96%,TT×DD-0五倍体诱导成功率为76.92%,TT×TT-0六倍体诱导成功率为12.50%。12月龄,各组合多倍体诱导成功率分别为91.43%(DD×PP-0,3n=74),60%(DD×DD-0,3n=75),85.19%(DD×TT-0,4n=100),87.50%(TT×PP-0,5n=124),83.33%(TT×DD-0,5n=125)和6.06%(TT×TT-0,6n=150)。12月龄各诱导组合多倍体倍性组成与1月龄相似,诱导成功率随诱导多倍体后代的倍性增加而降低。对不同诱导组合后代进行生长统计分析发现:不同诱导组合后代在生长早期差异不显著,生长后期表现出不同程度的差异。其中,以四倍体泥鳅为母本、大鳞副泥鳅为父本的诱导五倍体后代(TT×PP-0)12月龄时期全长和体质量值最大。同时,荧光受精细胞学观察证实冷休克处理通过抑制受精卵第二极体的释放获得三倍体型后代。

Six different polyploid loaches (DD×PP-0,3n=74;DD×DD-0,3n=75;DD×TT-0,4n=100;TT×PP-0,5n=124;TT×DD-0,5n=125 and TT×TT-0,6n=150.Female in the former) were induced using large scale loach ( Paramisgurnus dabryanus,PP),diploid (DD),and tetraploid (TT) dojo loach ( Misgurnus anguillicaudatus ) by cold shock.The cold shock was maintained for 30-35 min using 2-3 ℃ ice-water from 5 min after fertilization.The ploidy of induced progenies at different development stages were measured by flow cytometry.The results showed that the ratios of induced triploid progenies were 87.50% of DD×PP-0,53.85% of DD×DD-0,90.91% of DD×TT-0,84.96% of TT× PP-0,76.92% of TT×DD-0,and 12.5% of TT×TT-0 at 1 month of age.The ratios of induced polyploidy progenies were 91.43%(DD×PP-0,3n=74),60%(DD×DD-0,3n=75),85.19%(DD×TT-0,4n= 100),83.33%(TT×PP-0,5n=124),87.50%(TT×DD-0,5n=125) and 6.06%(TT×TT-0,6n=150) at 12 months of age,respectively.The ploidy composition of each group at 12 months was similar to that at 1 month of age.The ratio of induced polyploid progenies decreased with increasing ploidy levels.Growth performance of each group was measured and analyzed by statistical analysis.Little difference was observed at early development stage,while significant difference was observed at 12 months of age.The progenies of TT×PP-0 had the highest total length and body weight value at 12 months of age.Furthermore,the fertilization cytology observation confirmed that the second polar body was retained in fertilized ovum by cold shock to produce triploid-type progenies.