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Long pcr frustration and pron

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#1 Angeline



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Posted 29 September 2017 - 05:01 AM

I have once managed to amplified a 6.5 kb pcr.
However, the amplification is not reproducible again using the same pcr condition, cDNA samples, primers, high fidelity enzymes. Instead of getting the intended product, a band of less than 2kb base pair is obtained. I?ve tried reextracting the rna, redo reverse transcription, changing dntps, nucleuse free water, new aliquote of primers, new aliquote of enzyme and buffer, but still nothing seem to work. I appreciate All advices to reproduce the 6.5 kb product as it is crucial for my downstream. Thanks a million.

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