GAP-Seq: a method for identification of DNA palindromes
1 Gene Regulation and Chromosome Biology Laboratory, Frederick National Laboratory for Cancer Research, Cancer Research and Development Center, Frederick, MD 21702, USA
2 ABCC/ ISP, SAIC-Frederick, Inc., Frederick National Laboratory for Cancer Research, Frederick, MD 21702, USA
3 Department of Molecular Genetics, Cleveland Clinic Lerner Research Institute, Cleveland, Ohio 44195, USA
BMC Genomics 2014, 15:394 doi:10.1186/1471-2164-15-394Published: 22 May 2014
Closely spaced long inverted repeats, also known as DNA palindromes, can undergo intrastrand annealing to form DNA hairpins. The ability to form these hairpins results in genome instability, difficulties in maintaining clones in Escherichia coli and major problems for most DNA sequencing approaches. Because of their role in genomic instability and gene amplification in some human cancers, it is important to develop systematic approaches to detect and characterize DNA palindromes.
We developed a new protocol to identify palindromes that couples the S1 nuclease treated Cot0 DNA (GAPF) with high-throughput sequencing (GAP-Seq). Unlike earlier protocols, it does not involve restriction enzymatic digestion prior to DNA snap-back thereby preserving longer DNA sequences. It also indicates the location of the novel junction, which can then be recovered. Using MCF-7 breast cancer cell line as the proof-of-principle analysis, we have identified 35 palindrome candidates and physically characterized the top 5 candidates and their junctions. Because this protocol eliminates many of the false positives that plague earlier techniques, we have improved palindrome identification.
The GAP-Seq approach underscores the importance of developing new tools for identifying and characterizing palindromes, and provides a new strategy to systematically assess palindromes in genomes. It will be useful for studying human cancers and other diseases associated with palindromes.