2月19日，BMC Genomics發(fā)表了研究論文 “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”,。 該文章是由中國科學院水生生物研究所魚類系統發(fā)育學科組博士研究生童超波等人在何舜平研究員的指導下完成的,。
該文對常用的實驗方案進行了改進，首次將磁珠分選系統用于散在重復序列的分離,，取得了成功,。在鰱和鳙中分離得到了一個年輕的SINE家族（命名為HAmo SINE)，具有典型的SINE結構特征,。序列及結構分析表明,，該SINE家族拷貝間序列分歧小，為近期活躍復制增殖，其拷貝數通過熒光定量估計為10的5次方數量級,；分離得到了LINE家族(命名為HAmo LINE),，鑒定為其他魚類LINE2的同源物，屬于LINE2一支,。
該SINE家族和LINE家族具有一致的尾部序列和二級結構,，表明該SINE家族借助于其基因組內HAmo LINE編碼的反轉座酶系統實現自身近期的不斷增殖。經序列結構比較,，HAmo和Smal,，FokI家族雖然序列高度相似，但它們卻是在各自的進化譜系中依賴于同一起源的LINE家族,，獨立產生的。
該論文的亮點在于首次將磁珠分選系統應用于散在重復序列的分離,，代替了目前該研究領域傳統的通過構建文庫雜交掃描分離的方法,，這種新方法在實驗周期，成本,，效率等各方面明顯優(yōu)于傳統方法,，尤其對于傳統方法不能分離得到的低拷貝的SINE序列也能分離得到。其次,，分離得到的SINE家族和其他兩個在遠緣物種魚類內的SINE家族的高度相似性表現出自然界里面的罕見的獨立產生的SINE家族的趨同性,。
該學科組后續(xù)的研究將集中在利用已經建立SINE分離平臺，大規(guī)模分離SINE序列,，進行鯉科魚類系統發(fā)育和分子進化的研究,，并運用生物信息學在？科魚類中通過比較基因組學的方法篩選一大批潛在的候選SINE插入來研究其插入所揭示的系統發(fā)育意義,。（生物谷Bioon.com）
生物谷推薦原始出處：
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.
