Difference between revisions of "Primer Design for qPCR"
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# If you want to look at a specific region of the mRNA and an appropriate primer pair in that region is not selected, change the range under PCR template on the top | # If you want to look at a specific region of the mRNA and an appropriate primer pair in that region is not selected, change the range under PCR template on the top | ||
# Click '''Get Primers''' on the bottom and wait for results | # Click '''Get Primers''' on the bottom and wait for results | ||
| − | # Print this out, for future reference and when you order primers name them as follows species-Gene-seq.start-seq.end, for example '''mm-SREBF1-127-254'''. | + | # Print this out, for future reference and when you order primers name them as follows species-Gene-direction-seq.start-seq.end, for example '''mm-SREBF1-FWD-127-254'''. |
# Enter both primers in ExperimentDB | # Enter both primers in ExperimentDB | ||
Revision as of 13:52, 31 October 2013
Finding Known Primer Sets
- Check papers, especially supplementary tables
- Check PrimerBank http://pga.mgh.harvard.edu/primerbank/index.html
Making Your Own Primers From Entrez
- Find your RNA sequence on entrez by going to http://www.ncbi.nlm.nih.gov/nuccore (make sure its the correct species)
- Under analyse this sequence click Pick Primers
- Under PCR Product Size pick 70-150 as the range
- Under Exon/intron selection -> Intron Inclusion check the box by Primer pair must be separated by at least one intron on the corresponding genomic DNA
- If you want to look at a specific region of the mRNA and an appropriate primer pair in that region is not selected, change the range under PCR template on the top
- Click Get Primers on the bottom and wait for results
- Print this out, for future reference and when you order primers name them as follows species-Gene-direction-seq.start-seq.end, for example mm-SREBF1-FWD-127-254.
- Enter both primers in ExperimentDB
