Waters Paul D, Dobigny Gauthier, Waddell Peter J, Robinson Terence J
Research School of Biological Sciences, The Australian National University, Canberra, Australia.
Methods Mol Biol. 2008;422:227-37. doi: 10.1007/978-1-59745-581-7_14.
Long-interspersed nuclear element-1 (LINE-1) is a non-terminal repeat transposon that constitutes a major component of the mammalian genome. LINE-1 has a dynamic evolutionary history characterized by the rise, fall, and replacement of subfamilies. The distribution of LINE-1 elements can be viewed from a chromosomal perspective using fluorescence in-situ hybridization (FISH), as well as at the sequence level. We have designed LINE-1 primers from regions conserved among mouse, rat, rabbit, and human L1, which were able to amplify part of ORF2 from all eutherian (placental) mammals tested thus far. The product generated can be used as a FISH painting probe to examine the genomic distribution of L1 in different species. It can also be cloned and sequenced for phylogenetic analysis. Although FISH patterns resulting from LINE-1 chromosome painting and bioinformatic analyses have shown that this element accumulates in AT-rich regions of the genomes of mouse and human, our PCR amplified LINE-1 probe suggests that this is not a universal phenomenon, and that the patterns displayed in laurasiatherian, afrotherian and xenarthran species are less prominent. The "banding" like distribution of LINE-1 observed in human and mouse, therefore, appears to reflect aspects of genome architecture unique to Euarchontoglires (Supraprimates), the superordinal clade to which they belong. By sequencing the cloned amplicons used for FISH experiments and supplementing these with L1 sequences obtained from public databases, analysis by parsimony, distance-based, maximum likelihood, and "hierarchical Bayesian" or "marginal likelihood" methods provides a powerful adjunct to the FISH data. Using this approach, relatively intact LINE-1 from most placental orders tend to reflect accepted eutherian evolutionary relationships. This suggests that there were often only closely related copies active near branch points in the tree, that inactive copies tended to become extinct quite readily, and that for many orders recently active copies belong to a single lineage of this LINE.
长散在核元件1（LINE-1）是一种非末端重复转座子，是哺乳动物基因组的主要组成部分。LINE-1具有动态的进化历史，其特征是亚家族的兴起、衰落和更替。可以使用荧光原位杂交（FISH）从染色体角度以及在序列水平上观察LINE-1元件的分布。我们从小鼠、大鼠、兔子和人类L1中保守的区域设计了LINE-1引物，这些引物能够从迄今为止测试的所有真兽亚纲（胎盘）哺乳动物中扩增出部分ORF2。产生的产物可作为FISH涂染探针，用于检测不同物种中L1的基因组分布。它也可以被克隆和测序用于系统发育分析。尽管LINE-1染色体涂染和生物信息学分析产生的FISH模式表明该元件在小鼠和人类基因组的富含AT的区域中积累，但我们的PCR扩增的LINE-1探针表明这并非普遍现象，劳亚兽总目、非洲兽总目和异关节总目物种中显示的模式不太明显。因此，在人类和小鼠中观察到的LINE-1的“条带”样分布似乎反映了它们所属的超目分支真灵长总目（灵长总目）特有的基因组结构方面。通过对用于FISH实验的克隆扩增子进行测序，并补充从公共数据库获得的L1序列，采用简约法、基于距离法、最大似然法以及“层次贝叶斯”或“边际似然”方法进行分析，为FISH数据提供了有力的辅助。使用这种方法，大多数胎盘类目的相对完整LINE-1往往反映了公认的真兽类进化关系。这表明在进化树的分支点附近通常只有密切相关的拷贝活跃，不活跃的拷贝往往很容易灭绝，而且对于许多目来说，最近活跃的拷贝属于该LINE的单个谱系。
