DNA Sequencing and Gene Synthesis at PBI

The DNA Technologies Unit is a core DNA sequencing and DNA/gene/microarray synthesis facility, first established by PBI in 1988. With the development of microarray technologies and bioinformatics as key tools for the rapid discovery of genes expression and gene function, high-throughput DNA sequencing is an essential technology. The Unit provides high-throughput sequencing capabilities in support of agricultural genomics programs, PBI core projects, and collaborative projects. The Group has integrated in vitro enzymatic plasmid template preparation and high-throughput sequencing chemistry technologies with sequence resolution on a 96-capillary AB 3730xl DNA sequence analyzer. PBI has also acquired next generation sequencing platforms from Roche and Illumina that accommodate a wide range of genomics and functional genomics applications where increased throughput is desirable. Throughput capacity is 5760 sequencing runs per week with read lengths 800-1000 (Phred 20 score) on each unit.

The Unit not only supports PBI and collaborative projects but also provides DNA sequencing and DNA/Gene synthesis technology to universities, industry, and government departments.

Recently, PBI acquired the next generation sequencing system 454 Genome Sequencer FLX(tm) from Roche. With this system, the DNA sequencing will increase dramatically and the cost per base will decease significantly.

Services

Our DNA Technologies Unit offers a broad range of services:

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  High throughput colony picking, plasmid preparation and DNA sequencing of cDNA libraries (ESTs) and genomic sub-clone libraries; BAC-end sequencing and PCR product sequencing

• Gene synthesis, gene libraries and codon bias optimization

• Modified oligonucleotides (fluorescent, biotinylated, phosphorylated, modified bases)

Equipment

The laboratory houses:

• Three ABI 3730xl capillary electrophoresis DNA analyzers are focused on genomic projects and collaborations with a 96 and 384-well format for high-throughput sequencing.

• Roche 454 Genome Sequencer FLX™ instrument with Titanium chemistry capable of generating over 1 million 400bp reads or >500Mbp of sequence data per run and supporting many applications including de novo DNA sequencing of whole genomes or large insert clones (BACs), transcriptome profiling, SNP discovery, small RNA analysis, and metagenomics. http://www.454.com/
• Illumina Genome Analyzer IIx platform that can produce >150 million 2x75bp reads, or up to 25Gbp of data per run that will enable very deep coverage of samples at a relatively low cost. http://www.illumina.com/

• Bioautomation MerMade Model 192 DNA Synthesizer able to custom synthesize DNA molecules up to 150 bases long. The unit provides custom gene synthesis, gene libraries and codon bias adjustments as well as modified DNA including phosphorylated, biotinylated or fluorescent labeled oligonucleotides. Many other custom modifications are available on request. http://www.bioautomation.com/

• Two Dianex High Performance Liquid Chromatography (HPLC) systems enable custom purification of oligonucleotides and PCR products.

• A Combimatrix B3 microarray synthesizer capable of making custom 94K DNA microarrays. http://www.combimatrix.com/

Rates

These services are available to external organizations on a cost-recovery basis. Rates are dependent on the type of service available. Bulk rates for high-throughput plasmid preparation and DNA sequencing, oligonucleotide and gene synthesis are available.

Related Information

Institutes:

• NRC Plant Biotechnology Institute