BIOC 300D2 Lecture Notes - Lecture 9: Oligonucleotide, Rna-Seq, Sybr Green I
8 Jul 2016
School
Department
Course
Professor
Document Summary
Lecture 8: lab 11 week 2 mardi 15 mars 2016. Select amplicons that span intron-exon boundaries so genomic dna can be distinguished from cdna. : primers a and b is a good rt-pcr primer set, while primer c and d is a poor set. Quantification of mrna levels using real time pcr. Normal rt-pcr is only semi-quantitative because, in part, of the insensitivity of ethidium bromide. Thus real time pcr was developed because of: The need to quantitate differences in mrna expression. The availability of only small amounts of mrna (small amounts of tisuse, primary cells) Real time pcr is a kinetic approach in which you look at the reaction in the early stages while it is still linear. Measures fluorescence signals during the log linear phase of pcr. There is a straight line relationship between the amount of dna and cycle number when you look on a logarithmic scale. This is because pcr amplification is an exponential reaction.
