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S7806 Datasheet(PDF) 24 Page - List of Unclassifed Manufacturers |
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S7806 Datasheet(HTML) 24 Page - List of Unclassifed Manufacturers |
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24 / 32 page  20 V. TROUBLESHOOTING CpG WIZ™ p16 (S7800), p15 (S7802) and E-cadherin (S7804) Amplification Kits There is no visual evidence of products in any lane. Potential Problem: PCR amplification is not initiated. Recommendations: a. Confirm that all PCR components were added to the reaction tube. b. Confirm that the time and temperature settings on the thermocycler match those described in this manual. c. If performing "hot start" PCR using a "hot start" enzyme, verify the initial denaturation/activation time of 12 minutes at 95°C. d. For all other "hot start" methods, confirm the proper use of the reagents. e. Confirm that the PCR polymerase is still active. f. Confirm that no additional Mg2+ was added to the PCR reaction mix. g. The optimal annealing temperature is 60°C. If items #a-f have not remedied the problem, re-optimize annealing conditions to suit your amplification instrument. No amplification product is generated in the experimental samples using U, M and W primer sets, but products of the correct size are observed with the control samples. Potential Problem #1: Experimental DNA samples were degraded prior to chemical modification. Recommendation: Purify the genomic DNA again and repeat the chemical modification. Potential Problem #2: Chemically modified experimental DNA samples were stored for more than two months prior to PCR. Recommendation: Repeat the chemical modification on new genomic DNA samples. U or M primer sets are producing bands in all samples, including the "no DNA" controls. Potential Problem: PCR reagents are contaminated with amplification products. ? ? ? |
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