Quantitative PCR
Quantitative PCR (qPCR) is an accurate and sensitive method of quantifying the abundance of a target DNA sequence. This can be from genomic DNA or from cDNA resulting from the reverse-transcription of RNA. A polymerase chain reaction (PCR) is performed using sequence-specific primers and a DNA-binding dye or probe that provides a fluorescent signal proportional to the amount of product (or amplicon) formed during each amplification cycle. Analysis of the amplification curves allows samples to be quantified via a standard curve, or used to calculate relative expression levels between samples (eg, for validation of DNA arrays or confirming gene knockdown by siRNA). qPCR can also be used to analyse single nucleotide polymorphisms (SNPs), perform genotyping or assess methylation status.
Our Roche LightCycler 480 offers exceptionally quick run times (approx 1 h), high resolution melt curves and detection at six different emission wavelengths, making it suitable for use with a wide range of custom and commercially available dye chemistries, including SYBR Green I chemistry, hydrolysis and hybridisation probes. The LightCycler 480 also supports both 96 and 384 well formats, affording both flexibility and capacity.
In addition to the classical uses of qPCR, genotyping by known SNP analysis can be performed on the LightCycler 480, using hydrolysis probes (eg, Applied Biosystems' SNP Genotyping Assays), hybridisation probes or high resolution melt (HRM) curve analysis. HRM analysis can also be used to screen regions of DNA for unknown variation or to quantify the degree of methylation of selected sequences.
We're happy to discuss your particular needs and help advise your experimental design, from sample preparation to data analysis.
Contacts:
Dr Alun Hughes (Scientific and application enquiries)
Email: a.hughes@abdn.ac.uk, Ext.55162
Mrs Lorna Smith (Customer and billing enquiries)
Email: l.m.smith@abdn.ac.uk, Ext.55930