cDNA conversion for RT-PCR application

June 02, 2015

University of Nebraska Medical Center
Pharmacology and Experimental Neuroscience
Graduate Student

Overall

Quality of Results

Ease-of-Optimization

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Company:

Life Technologies

Product Name:

RevertAid First Strand cDNA Synthesis Kit

Catalog Number:

K1621

The aim of my study was to determine gene changes associated with T cell profiles using RT-PCR. Specifically, I wanted to use this product to convert mRNA to cDNA for use in my PCR application. This kit allowed me to easily do so.

Experimental Design and Results Summary

Application

cDNA synthesis

Starting Material

mRNA isolated from CD4+ T cells

Protocol Overview

I isolated fresh CD4+ T cells from spleen and lymph nodes of drug-treated mice. I then isolated RNA and used this kit to convert mRNA into cDNA for future use in RT-PCR.

Tips

Says it will yield 20 ul per cDNA reaction, but sometimes I did not get this much

Results Summary

I was able to successfully and efficiently convert mRNA to cDNA for PCR application. However, on a rare occasions, conversion did not work successfully so I had to redo it.

Additional Notes

None

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Summary

The Good

Easy to use and allows for successful conversion from RNA to cDNA

The Bad

Each reaction yields a small amount (20 ul) so for 96 well plates, you must undergo multiple reactions in order to get enough cDNA

The Bottom Line

Overall, the conversion kit was easy to use and gave fairly good results. However, sometimes the conversion was not successful and needed to be redone. Because of this, if you don't have a ton of starting material (mRNA), this kit may not be the best choice.

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