This laboratory exercise is intended for those who have a fundamental background in PCR.
Many types of online primer design software are available, which can be used free of charge to design desirable SYBR® Green-based qPCR primers. As SYBR® Green dye binds to any double-stranded DNA product, its success depends greatly on proper primer design. SYBR® Green is very simple to use and cost efficient. The four most commonly used fluorescent chemistries are SYBR® Green dyes and TaqMan®, Molecular Beacon or Scorpion probes. The process uses reverse transcription polymerase chain reaction (RT-PCR), coupled with fluorescent chemistry, to measure variations in transcriptome levels between samples. Real-time PCR (quantitative PCR or qPCR) has become the preferred method for validating results obtained from assays which measure gene expression profiles.
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