CRISPR gene cuts may offer new way to chart human genome

Previous studies have shown that a decrease in DNA methylation in KRT19 is associated with tumor spread.

In the breast cancer cell lines they studied, the Johns Hopkins team was able to generate an average of 400 "reads" per basepair, a reading "depth" hundreds of times better than some conventional sequencing tools.

Among their samples of human breast cancer tumor tissue taken at biopsies, the team was able to produce an average of 100 reads per region.

"This is certainly less than what we can do with cell lines, but we have to be more gentle with DNA from human tissue samples because it's been frozen and thawed several times," says Timp.

In addition to their studies of DNA methylation and small mutations, Timp and Gilpatrick sequenced the gene commonly associated with breast cancer: BRCA1, which spans a region on the genome more than 80,000 bases long.

"This gene is really long, and we were able to collect sequencing reads which went all the way through this large and complex region," says Gilpatrick.

"Because we can use this technique to sequence really long genes, we may be able to catch big missing blocks of DNA we wouldn't be able to find with more conventional sequencing tools," says Timp.

In addition to its potential to guide treatment for patients, Timp says the combination of CRISPR technology and nanopore sequencing provides such depth that it may help scientists find new disease-linked gene alterations specific to one allele (inherited from one parent) and not another.

Timp and Gilpatrick plan to continue refining the CRISPR/nanopore sequencing technique and testing its capabilities in other tumor types.

Story Source:

Materials provided by Johns Hopkins Medicine.