RNA was isloted with and without RNAlater. Although the yeild was reduced (190 µg v. 240 µg), the quality of the RNA seems to be improved ize="5">
Use 3DNA Capture Sequence for Dowstream PCR Primer
In conjunction with the atp15 5' primer and in the absence of cDNA, the Cy3-capture 3' primer produced PCR artifact bands but the Cy5 primer did not (green arrow). Since the cDNA was produced with a Cy5 capture sequence downstream of a poly-dT tail, a PCR product using atp15 and Cy5 primers should produce a smear up to approximately 380 base pairs. Therefore, it appears that this combination was able to detect the successful production of cDNA. 20% of the cDNA was consumed in this PCR.
It is worth noting that the 4plex primers without cDNA produced the ~179 band seen on the previous gel.