GenCh13answers - The Dynamic Genome: Transposable Elements...

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The Dynamic Genome: Transposable Elements 1. Mutations in gal can be generated, and from these strains, λ dgal phage isolated. Through hybridization of denatured λ dgal DNA containing the mutation with wild-type λ dgal DNA, some of the molecules will be heteroduplexes between one mutant and one wild-type strand. If the mutation was caused by an insertion, the heteroduplexes will show a “looped out” section of single-stranded DNA, confirming that one DNA strand contains a sequence of DNA not present in the other. The text also illustrates a method to compare the densities of gal + -carrying λ phage with gal -carrying phage. In this experiment, the gal -phage are denser, indicating that they contain a larger DNA molecule. If the gal genes are cloned, direct comparison of the restriction maps or even the DNA sequence of mutants compared with wild type will give specific information about whether any are the results of insertions. Using primers that flank the gene, PCR can be used to determine the size of the intervening fragment. Insertions would result in a larger-sized fragment compared to wild type. 2. In replicative transposition, transposable elements move to a new location by replicating into the target DNA, leaving behind a copy of the transposable element at the original site. If, on the other hand, the transposable element excises from its original position and inserts into a new position, this is called conservative
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GenCh13answers - The Dynamic Genome: Transposable Elements...

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