DNA sequencing at the John Innes Centre

JIC and the Sainsbury Laboratory share a sequencing service that integrates an ABI3730 high throughput sequencer and liquid handling robotics into a single cohesive site-wide service. Three staff manage the service, which currently delivers over 1,000 sequencing reads and around 500 fingerprint (gene scan) mapping reads per week.

The service had a significant role in the recent international Arabidopsis genome sequencing project. Currently the service is engaged on several BBSRC and EC funded functional genomics projects, including the sequencing of transposon insertion sites and Arabidopsis ecotypes.

A major project is the high throughput fingerprint (gene-scan) mapping of the wheat genome in which specific regions of the wheat genome will be sequenced. Interesting regions will be identified using genetic mapping information from wheat and synteny data from the rice genome sequence. Targets include the Ph1 chromosome pairing locus and disease resistance genes. Regions of Lotus japonicus, pea and Medicago truncatula genome will be sequenced, based on mapping of interesting mutant alleles. Other projects include sequencing the genome of the cereal pathogen Erisyphe.

A gene disruption programme in Streptomyces will generate over 15,000 mutants many of which will be checked by sequencing. In addition several projects exploring gene induction (e.g. by bacterial, viral or fungal pathogens or by abiotic stress) will use EST sequencing to identify regulated genes from cDNA-AFLP or subtractive libraries.

The service also provides high throughput of small-scale sequencing projects required to check plasmid constructs and for gene identification.

Impending improvements include the robotic preparation of DNA templates and sequencing reactions, to further increase throughput and decrease unit costs.