PCR Flashcards
What does reverse transcription polymerase chain reaction do ?
Technique used to detect and quantify mRNA
What does reverse transcription polymerase reaction work in terms of genes ?
Shows if a specific gene is being expressed in a given cell type
What is the advantage of RT-PCR over northern blot and RNAase protection assay ?
Can be used to quantify mRNA levels from smaller samples
What happens first in RT-PCR ?
An RNA strand is first reverse transcribed into its DNA complement (cDNA)
How is cDNA amplified ?
Using end point or real time PCR
What helps the transcription of mRNA to cDNA ?
- Reverse transcriptase
2. Oligo dT primer
What do oligo dT primers do ?
Bind to the polyA tail of mRNA
What can DNA polymerase only act on ?
DNA templates
What happens if mRNA is present (step 1) ?
Reverse transcriptase and primer will anneal to the mRNA sequence and transcribe a cDNA
(Step 2) What is cDNA replicated with ?
Primers and Taq DNA polymerase
What are the PCR products ?
DNA strands
What are reporter genes used for ?
To monitor various aspects of gene expression in model organisms
What does the reporter gene act as a surrogate for ?
Original gene
What is the reporter gene subjected to ?
The regulatory properties of the attached promoters
What are transcriptional gene fusions used for ?
Identify many cis-acting regulatory elements that confer tissue specific and developmental specific patterns of gene expression in plants and animals
