High-throughput screening: Genotyping by PCR
When is conventional PCR appropriate as a high-throughput screening method?
Conventional endpoint PCR is an effective, proven, and affordable method for high-throughput screening projects. In these situations, conventional PCR is performed on large numbers of samples and reactions are analyzed by agarose gel electrophoresis. Applications may include genotyping by PCR or doing routine, two-step semi-quantitative RT-PCR screening for expression of marker genes.
How can an efficient workflow help ensure success with high-throughput PCR screening projects?
When performing dozens, hundreds, or even thousands of PCR reactions, efficiency is critical: every improvement in workflow efficiency leads to significant savings in time. Eliminating pipetting steps reduces potential for contamination and error, as well as minimizes repetitive strain-induced injury. Simplifying the screening procedure also enables better standardization between different laboratory personnel, which can be crucial as screening projects scale up.
Typical endpoint PCR screening assays include:
- determining presence or absence of a target sequence (e.g., a transgene)
- allelic determination (e.g., variable repeats, RFLP analysis, insertions, deletions)
- identifying mutants during forward or reverse genetic screens
- screening cell lines for expression of biomarker genes by RT-PCR
EmeraldAmp GT PCR Master Mix: Optimized for High-Throughput PCR
Optimized to enable efficient workflow during routine endpoint PCR as well as PCR screening projects such as genotyping large numbers of samples, Takara’s EmeraldAmp GT PCR Master Mix provides excellent performance at an affordable price. Assemble reactions simply by adding primers and template to the 2X premix, which contains buffer, PCR polymerase, dNTPs, gel loading dyes, and a density reagent. After PCR, samples can be loaded directly onto an agarose gel for electrophoresis, used for restriction digestion, or easily cloned into a T-vector. EmeraldAmp GT PCR Master Mix is suitable for amplifying genomic DNA products up to 5 kb, even for GT- or AT-rich templates.