Figure 1

Techniques for identifying transposon insertions. (A) Polymerase chain reaction (PCR)-based assays detect transposable element (TE) insertions. L1 display utilizes primers specific to particular subfamilies of LINE-1 elements. Using this method, candidate dimorphic L1 insertions have been identified. The ATLAS technique employs the principles of L1 display and suppression PCR. Genomic DNA is digested and ligated to oligonucleotide primers, and used as a template in a PCR reaction containing L1 and linker-specific primers. Primary PCR products are then used as templates in a linear PCR reaction containing a radiolabelled subfamily-specific L1 primer. Radiolabelled products are detected by electrophoresis and autoradiography. (B) A comparative genomics approach to identify TE insertions and deletions is depicted. For example, the completion of the draft chimpanzee genome sequence provided an opportunity to identify recently mobilized transposons in humans and chimpanzees. If a transposon insertion is present in only one of the two genomes, it is inferred that the insertion occurred since the existence of their most common recent ancestor (~6 million years ago). (C) A paired-end mapping approach is shown. This method entails generating paired-ends of several kilobase fragments, which are sequenced using next generation sequencing methodologies. Differences between the paired-end reads and a reference genomic region reveal the presence of structural variation. Simple insertions and deletions can be detected using this method. (D) A next generation sequencing method is shown. Selective amplification of the 3' end of a transposon is performed followed by deep sequencing. This short-read sequencing approach is able to detect precise insertion positions. (E) Microrray-based methods involve the hybridization of ligation-mediated PCR products to genomic tiling arrays. Specifically, vectorettes are ligated to restriction enzyme-digested genomic DNA. The amplified fragments include the 3' end of a transposon sequence and unique flanking genomic DNA. These amplicons are hybridized to genomic tiling microarrays.