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Department of Plant Pathology

DNA Sequencing and Genotyping Facility

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DNA Sequencing

We use an Applied Biosystems 3730 DNA Analyzer for sequencing. It has the capacity to analyze 96 samples every 4 hrs and get 750-900 bases per sample. That’s over 48,000 bases of DNA per run. It is fully automated and can analyze 4-96 well plates without human intervention. At full capacity, it can obtain more than 300,000 bases of DNA per day.

Typically, customers will provide us with DNA samples which we use as templates for cycle sequencing. We use Applied Biosystems Big Dye chemistries for sequencing. Also, we run samples that customers have themselves performed cycle sequencing. The customer receives results in the form of chromatograms and text files.

The 3730 is very different and less forgiving than other old generation ABI machines used by other facilities. Before you submit samples, you must read Sample Prep Info.

DNA Fragment Analysis

Applied Biosystems 3730 DNA analyzer can provide excellent resolution on molecular marker analysis such as microsatellite, AFLP, SNP, trAP etc. It can resolve 1 bp nucleotide difference between two genotypes. Also the 3730 provides high-throughput multiplex fragment analysis and has the capacity to analyze 384 samples per microsatellite marker every 2-3 hrs at reasonable low cost.

For genotyping, customers will provide us with ready to run PCR products and we will run them in the ABI 3730. The chromatograms data will be sent to customers in a CD. Data can be scored using specific software in the facility. Different from sequencing, genotyping work usually uses many different primers and has a large number of samples to be analyzed per experiment. Customers will use their own primers for PCR. Staff in the facility can help customers in planning experiments, optimization of PCR protocol and data scoring.