User:Saroj Pandey/Notebook/SNP PCR optimization/2014/09/25: Difference between revisions
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* A PCR using 4 primers (@ outer and 2 inner primers) was performed | * A PCR using 4 primers (@ outer and 2 inner primers) was performed | ||
* Forward inner primer ends in G and reverse inner primer ends in C | * Forward inner primer ends in G and reverse inner primer ends in C | ||
* Expected band sizes for taster: 992bp and 510bp | * Expected band sizes for taster: 992bp and 510bp | ||
* Expected band sizes for taster: 992bp and 522bp | * Expected band sizes for taster: 992bp and 522bp | ||
[[Image:multiplexPCR.JPG]] | [[Image:multiplexPCR.JPG]] | ||
[[Image:multiplexPCR_26.09.jpg|right|thumb|350px|Electrophoresis]] | [[Image:multiplexPCR_26.09.jpg|right|thumb|350px|Electrophoresis]] | ||
Observation: | Observation: | ||
• No bands are seen at 992bp which tells that amplification with the outer primers was not successful | • No bands are seen at 992bp which tells that amplification with the outer primers was not successful | ||
• Faint bands are seen at about 500bp in both the taster and the non-taster, but | • Faint bands are seen at about 500bp in both the taster and the non-taster, but | ||
Conclusion: | Conclusion: | ||
• Since the expected products are of similar lengths, this PCR is unable to distinguish between the two variants and other set of primers could be used | • Since the expected products are of similar lengths, this PCR is unable to distinguish between the two variants and other set of primers could be used | ||
Revision as of 03:59, 1 October 2014
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Allele Specific PCR and Multiplex PCR
• No bands are seen at 992bp which tells that amplification with the outer primers was not successful • Faint bands are seen at about 500bp in both the taster and the non-taster, but
• Since the expected products are of similar lengths, this PCR is unable to distinguish between the two variants and other set of primers could be used
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