RGCB has a centralized Genomics facility to meet the demand of scientists in various research fields. This facility is greatly helping their research work, catering to the sequencing needs of the Institute. Services include DNA sequencing, Genotyping using DNA analyzers, Real time PCR and Droplet digital PCR
Automated DNA Sequencing Facility is equipped with 3730 DNA Analyzer (48 capillary high throughput DNA Sequencer) and 3730xL DNA Analyzer (96 capillary high throughput DNA Sequencer). The fluorescence-based DNA sequencing systems, with capillary electrophoresis is having 96 capillaries operating in parallel for Sanger sequencing. The system automatically performs sample injection, gel matrix replacement, DNA separation, detection, and data analysis. This system sequence 48-96 samples in 2 hours' time and perform up to 12 runs per day. Average read length with big dye terminator (BDT) chemistry is 800bp. This system can also be used for fragment analysis, AFLP, genotyping and SNP analysis.
The facility equipped with RealTime PCRs (7900HT Real-Time, 96-384 well format and 7500 Real time PCR-96 well) and droplet digital PCR (QX 100-Biorad). The systems can be used for methods such as
The Core accepts DNA samples (pure plasmids, mini-prep plasmids, M13 clones, cosmids, lambda clones, PCR products, gel isolated fragments, bacterial genomic), sequenced PCR products, and processes them according to protocols for Applied Biosystems DNA Sequencers (BDT Chemistry). One sequencing run (one template, one primer) typically produces 700-800 nucleotides of high-quality sequence data or more for good quality template and a well-designed primer. The results are returned in three forms: a raw sequence file, sequence with blue score and a chromatogram file. The sequence file (a flat ASCII text file) is returned to the researcher by electronic mail.
Each sequencing plate includes at least one lane of standard DNA known to generate good sequence. The sample handling and data analysis are extremely reliable and have a very low rate of error. If any problems are indicated in our sample processing, we repeat the affected samples on request. We typically process samples within two days after receipt. Samples are processed Monday through Friday for overnight instrument runs. The data are processed and returned by email to the users.
Manoj P, PhD
Asst General Manager (Tech services)