Questions pertaining to the synthesis of DNA from RNA.

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8
votes
2answers
74 views

Is there a Reverse Transcription optimization for long, 9kb, transcripts?

Has anyone optimized RT for long transcripts (9kb)? The downstream application will be PCR amplification and Illumina library prep. It will be trivial to make internal primers sets for the PCR that ...
6
votes
2answers
631 views

Reverse transcription PCR optimization

What is the ideal amount of RNA to use for the RT? and how much cDNA to use then for the PCR? I did RT with a solution of RNA of 0.36 ug/ul. Then for my PCR I used 1 ul of the cDNA obtained and used ...
3
votes
2answers
32 views

Measuring reverse transcription yield?

Is there any measurement I could perform after RT that allows me to check the efficiency that the procedure had? Nanodrop cannot be used as remnants of RNA and poly T primers mask measure, or?
3
votes
0answers
47 views

Large scale reverse transcription?

I need to make RNA:DNA duplexes. I can make 100 to 200 ug of mRNA through in vitro transcription, and I know how to use reverse transcription to make a cDNA library, but I have questions with this. ...
2
votes
1answer
27 views

HIV-RT nucleotide contact residues

I'm investigating HIV-RT 3KLF and 1LWF trying to find a perfect pose of thymine. Which residues on HIV-RT is Thymine supposed to contact on 3KLF and 1LWF? Where exactly is the nucleotide binding ...
0
votes
1answer
61 views

qrt-pcr and short fragments

I plan to measure the effects of Tenofovir a Nucleotide analog reverse-transcriptase inhibitors useing qrt-pcr with HIV-RT as the rt enzyme. Tenofovir causes early termination of the reverse ...