Study On The Genetic Diversity Of Pelteobagrus Fuluidraco And Silurus Asotus In Henan By ISSR Markers

Genetic diversity was one of the core in the conservation biology,which refers to the level of genetic variation between various groups in one population and the degree of variation of different individuals in the same group.ISSR molecular markers technology could be called a preferable method to explore the genetic diversity,on account of the advantage of speediness,efficiency and sensitiveness.In this research,ISSR molecular markers technology was devoted to make a comprehensive analysis of genetic diversity and genetic structure in Pelteobagrus fulvidraco and Silurus asotus,which belongs to Siluriformes.The work for Pelteobagrus fulvidraco and Silurus asotus in Henan has significant development in protection of species resources,sustainable development and cultivation of population.Muscular tissue of Pelteobagrus fulvidraco and Silurus asotus were conserved in 95% ethyl alcohol,they could be preprocessed with four methods and then compared the consequence of genomic DNA extraction.The results showed that the pretreatment sample of STE buffer was the desired pre-treatment to extract muscular tissue in ethyl alcohol,because after pretreatment of STE buffer,the bands of DNA were clear,orderliness and out of degradation,in the meantime,the values of A260/A280 were berween 1.8-2.0.Fifteen ISSR primer were respectively singled out by literature and consult relatives to discuss the level of gene in Pelteobagrus fulvidraco and Silurus.The orthogonal test design method and re-optimization in Pelteobagrus fulvidraco and Silurus asotus,which were applied to establish optimal of ISSR-PCR reaction condition.The 25 μL system in Pelteobagrus fulvidraco and Silurus asotus included 45 ng DNA template,1.5mmol/L Mg²⁺,0.4mmol/L dNTPs,0.4μmol/L primer,0.5U Taq DNA polymerase.Not only that,the optimal procedure of ISSR-PCR was also determined,including initial denaturation at 94℃ for 5min,unwinding at 94℃ for 30 s,thermal denaturation at 45⁶0℃ for 40 s,prolonged at 72℃ for 90 s,re-prolonged at 72 for 5min,conserved at 4℃.Simultaneously cycle index and annealing temperature with primers were optimized and the ideal cycle was 40.In the aspect of genetic diversity,Pelteobagrus fulvidraco in three populations were analyzed,128 sites were emerged by amplification and the polymorphic loci were 54,the proportion was 42.19%.Thegenetic diversity parameters of three population included that PPB were 28.91%,27.34% and 28.91%,Na were 1.2891,1.2734 and 1.2891,Ne were 1.1795,1.1758 and 1.1985,h were 0.1032,0.1022 and 0.1130,I were 0.1536,0.1515 and 0.1657.As shown above,genetic distribution of Pelteobagrus fulvidraco in Henan kept at a high level and was in the order of DJHS > QHHS > NWHS.The index of genetic divergence was high,which based on Gst of 0.3510.Shannon indices indicated that the variation of Pelteobagrus fulvidraco include 34.03% in intrapopulation and 65.97% in interpopulation.Consequently,the heritable variation of Pelteobagrus fulvidraco derived from both sides and greater towards interpopulation.The gene flow was lower which based on Nm of 0.9247.It could be caused by genetic drift or the effect of natural selection greater than gene exchange,and then devote to genetic differentiation.In the aspect of genetic diversity,Silurus asotus in three populations were analyzed,130 sites were emerged by amplification and the polymorphic loci were 59,the proportion was 45.38%.The genetic diversity parameters of three populations included that PPB were 39.23%,40.00% and 34.62%,Na were1.3923,1.4000 and 1.3462,Ne were 1.3232,1.3034 and 1.2667,h were 0.1742,0.1669 and 0.1465,I were0.2487,0.2411 and 0.2112.The genetic distribution index of Silurus asotus between three area were higher and was in the order of NWNY > QHNY > DJNY.The index of genetic divergence was high,which based on Gst of 0.1820.Shannon indices indicated that the variation of Silurus asotus include 17.27% in intrapopulation and 82.73% in interpopulation.The gene flow was more frequently than other populations and the level of genetic differentiation between populations was not high,which based on Nm of 2.2470.The reasons for variation could be caused by fusion of gene flow or the uniform selection pressure.