Note+about+reverse+transcriptase+of+In-Fusion+SMARTer+Directional+cDNA+Library+Construction+Kit+1-14-13

I noticed that many of the products from the kit were 5' truncated (did not contain the 5' end of the genes). Perhaps the reverse transcriptase did not make full length cDNA .

here's some information from a Clontech technical service email https://mail.google.com/mail/u/0/?ui=2&shva=1#label/Career/13c3aa3becc4b474

Note that PrimeScript™ Reverse Transcriptase may work better than SMARTScribe™ Reverse Transcriptase on GC-rich templates.

To alleviate formation of RNA secondary structure you may also try the following modification of the SMART cDNA Synthesis Protocol:

· use longer RNA denaturation time - 72oC for 5 min

· cool on ice for about 2 min

· briefly spin RNA solution by centrifugation - 10 seconds.

· quickly add Master Mix - proceed further according to the RACE Protocol