商城肥鲵二碱基重复和四碱基重复微卫星DNA的结构特征及对筛选效率的影响

两栖类有尾目物种的微卫星分离中的筛选成功率常常较低。为探索微卫星结构对筛选效率的影响,本研究通过AFLP快速分离法(fast isolation by AFLP of sequences containing repeats,FIASCO)对商城肥鲵(Pachyhynobius shangchengensis)二碱基重复类型和四碱基重复类型微卫星进行分离,并对微卫星序列进行了分析。研究中发现二碱基微卫星位点多以微卫星DNA家族形式存在,并因此导致了微卫星位点分离较低的筛选率;在四碱基重复的微卫星位点中未发现微卫星DNA家族的存在。对研究中得到的3个微卫星DNA家族的分析发现,同一家族的上、下游侧翼序列变异程度存在差异;毗邻微卫星重复单元区的侧翼序列碱基变异程度较高,而较远处的区段则相对保守。这些结构特征可能反映出微卫星DNA家族在演化中的复杂性。本文的研究结果提示在两栖动物的一些类群中,微卫星的筛选必须考虑微卫星DNA家族的影响,选取适宜的碱基重复类型将是决定筛选效率的关键。

Structural characteristics of di-nucleotide/tetra-nucleotide repeat microsatellite DNA in Pachyhynobius shangchengensis genomes and its effect on isolation

A low success ratio is often encountered in the microsatellite isolation from the species of Caudata.In the present study,di-nucleotide/tetra-nucleotide repeat microsatellite loci were isolated following the FLASCO(fast isolation by AFLP of sequences containing repeats) approach and the sequences were ana-lyzed for the purpose of learning how microsatellite structure impacted on isolation efficiency.We found that most di-nucleotide repeat microsatellite loci were involved in the microsatellite DNA family,which resulted in low isolation efficiency.In contrast,no microsatellite DNA family members were found in the tetra-nucleotide repeat microsatellite loci.On the basis of analysis of the three microsatellite DNA families found in the present study,there were significant differences in the extent of variation between the upstream and downstream flanking sequences in same microsatellite DNA family.This variation was the highest in the sequence sections adjacent to the core repeated units,but more conservative in other regions,which might re-flect the complexity of the microsatellite DNA family evolution.These results suggest that in the process of microsatellite development for Caudata,we must pay attention to interference from microsatellite DNA fam-ily evolution.The choice of appropriate nucleotide repeats is key to obtaining a high isolation efficiency.