2月19日，BMC Genomics发表了研究论文  “Bead-probe complex capture a couple of SINE and LINE family from genomes of two closely related species of East Asian cyprinid directly using magnetic separation”。  该文章是由中国科学院水生生物研究所鱼类系统发育学科组博士研究生童超波等人在何舜平研究员的指导下完成的。

该文对常用的实验方案进行了改进，首次将磁珠分选系统用于散在重复序列的分离，取得了成功。在鲢和鳙中分离得到了一个年轻的SINE家族（命名为HAmo SINE)，具有典型的SINE结构特征。序列及结构分析表明，该SINE家族拷贝间序列分歧小，为近期活跃复制增殖，其拷贝数通过荧光定量估计为10的5次方数量级；分离得到了LINE家族(命名为HAmo  LINE)，鉴定为其他鱼类LINE2的同源物，属于LINE2一支。

该SINE家族和LINE家族具有一致的尾部序列和二级结构，表明该SINE家族借助于其基因组内HAmo LINE编码的反转座酶系统实现自身近期的不断增殖。经序列结构比较，HAmo和Smal，FokI家族虽然序列高度相似，但它们却是在各自的进化谱系中依赖于同一起源的LINE家族，独立产生的。

该论文的亮点在于首次将磁珠分选系统应用于散在重复序列的分离，代替了目前该研究领域传统的通过构建文库杂交扫描分离的方法，这种新方法在实验周期，成本，效率等各方面明显优于传统方法，尤其对于传统方法不能分离得到的低拷贝的SINE序列也能分离得到。其次，分离得到的SINE家族和其他两个在远缘物种鱼类内的SINE家族的高度相似性表现出自然界里面的罕见的独立产生的SINE家族的趋同性。

该学科组后续的研究将集中在利用已经建立SINE分离平台，大规模分离SINE序列，进行鲤科鱼类系统发育和分子进化的研究，并运用生物信息学在？科鱼类中通过比较基因组学的方法筛选一大批潜在的候选SINE插入来研究其插入所揭示的系统发育意义。

原始出处：

BMC Genomics 2009, 10:83doi:10.1186/1471-2164-10-83

Bead-probe complex capture a couple of SINE and LINE family from genomes of two closely related species of East Asian cyprinid directly using magnetic separation

Chaobo Tong , Baocheng Guo  and Shunping He

Abstract (provisional)

Background

Short and long interspersed elements (SINEs and LINEs, respectively), are two types of retroposons that are active in shaping the architecture of genomes and are powerful tools for studies of phylogeny and population biology. Here we developed a special protocol to apply a biotin-streptavidin bead system to isolate interspersed repeated sequences rapidly and efficiently, in which SINEs and LINEs were captured directly from digested genomic DNA by hybridization to a bead-probe complex in solution. This method was used instead of the traditional strategy using genomic library construction and screening.

Results

New SINEs and LINEs that shared an almost identical 3'tail were isolated and characterized in silver carp and bighead carp of two closely related species. These SINEs (34 members), designated the HAmo SINE family, showed little divergence in their sequences and were flanked by obvious TSD, indicating that HAmo SINE is a very young family. The copy numbers of this family was estimated to 2x10(5) and 1.7x10(5) per haploid genome by Real-Time qPCR, respectively. The LINEs, identified as homologs of LINE2 in other fishes, had a conserved primary sequence and secondary structures of the 3'tail region that was almost identical to that of HAmo SINE. This evidence suggests that HAmo SINEs are active and were amplified recently, utilizing the enzymatic machinery for retroposition of HAmoL2 through the recognition of higher-order structures of the conserved 42-tail region. We analyzed the possible structures of HAmo SINE that lead to successful amplification in the genome and deduced that HAmo SINE, SmaI SINE and FokI SINE shared similar sequences and were probably generated independently and created by the LINE family within the same lineage of a LINE phylogeny in the genomes of different hosts.

Conclusion

The results show the advantage of the novel method for retroposons isolation and a pair of young SINE family and its partner LINE family in two carp fishes, which strengthened the hypotheses of the slippage model for initiation of reverse transcription, retropositional parasitism of SINEs on LINEs, the formation of the stem loop structure in 3'tail region of some SINEs and LINEs and the mechanism of template switching in generating new SINE family.