Sequencing the unsequenceable: Genome Center technology advances Fragile X research

The UC Davis Genome Center has been applying one of the world’s most advanced commercial genome sequencing machines to study the developmental disorder, Fragile X syndrome.

The DNA code is made up of four letters, A, T, C and G. Fragile X syndrome occurs when a large number of three-letter repeats CGG appears in part of the fragile X mental retardation gene, FMR1. A run of more than 200 repeats causes full-blown fragile X syndrome, switching off the gene altogether and causing serious intellectual disability. But shorter sequences of repeats can also cause a range of problems including learning disabilities, seizures and fragile X-associated tremor/ataxia syndrome.

Paul Hagerman, a professor of biochemistry and molecular medicine at the
UC Davis School of Medicine, studies these disorders related to the CGG repeats in the FMR1 gene. Most individuals with fragile X actually have a mix of FMR1 genes with different lengths of repeats, and that can affect their condition, because some of their cells may be able to make the FMR1 protein at least partially.

Hagerman wanted to study the different lengths of repeats in patients and see how they are related to different conditions. However, sequencing such long pieces of DNA individually is not possible with most current technology.

Hagerman partnered with the UC Davis Genome Center to use an advanced sequencing machine built by Pacific Biosciences. The machine reads a single DNA molecule at a time, and can “read” a much longer stretch of DNA in one run than conventional DNA sequencing techniques.

Using the single-molecule approach, they were able to sequence the full range of length repeats, up to 750 base-pairs. The approach also gives information about whether DNA is chemically altered by methylation, which regulates gene activity.

The ability to sequence the fragile X genes and look at their size and and methylation status is going to be greatly helped by single-molecule sequencing, Hagerman told the trade publication GenomeWeb.

The same approach could also be applied to other disorders where the length of a repetitive stretch of DNA is important, such as Huntington’s Disease, Hagerman and colleagues wrote in their paper.

A paper describing the work is published online in the journal Genome Research.



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