The formation of DNA from an RNA template using reverse transcription leads to the formation of double-stranded complementary DNA or cDNA. The challenges with this process include 1. Maintaining the integrity of RNA, 2. Hairpin loops or other secondary structures formed by single-stranded RNA can also affect cDNA synthesis, and 3. DNA-RNA hybrids, which may result when the first strand of cDNA is formed. For the first challenge, using workflows that involve proper isolation and storage of RNA, and maintaining a nuclease-free environment helps obtain RNA with ideal 260/230 ratios. Using a reverse transcriptase that can tolerate high temperatures (50-55oC), overcomes obstacles imposed by secondary RNA structures. Finally, RNase H has the ability to hydrolyze RNA before the formation of a second cDNA strand. It is important to ensure that RNase H activity is optimal because higher RNase H activity leads to premature degradation of the RNA template. Many reverse transcriptases offer built-in RNase H activity.
Get tips on using DMEM/F-12 to perform 3D Cell Culture Media Mouse primary breast ephitelial cells-Mammospheres
Get tips on using DMEM/F-12 to perform 3D Cell Culture Media Mouse primary lung epithelial cells-organoids
Get tips on using DMEM/F-12 to perform 3D Cell Culture Media Mouse primary mammary ephitelial cells- organoids
Get tips on using Gibco™ RPMI 1640 Medium to perform Stem cell culture media Mouse trophoblast stem cells
Get tips on using Gibco™DMEM/F-12 to perform Stem cell culture media Mouse trophoblast stem cells
Get tips on using Gibco™DMEM/F-12 to perform Stem cell culture media NCH421K cells primary glioma
Get tips on using LC3A/B (D3U4C) XP® Rabbit mAb #12741 to perform Autophagy assay cell type - Mel cells
Get tips on using Dulbecco’s Modified Eagle’s Medium (DMEM) (1X),liquid to perform Mammalian cell culture media HSG cells
Get tips on using NucleoSpin® RNA to perform RNA isolation / purification Cells - immortalized OVCAR-3
Fill out your contact details and receive price quotes in your Inbox
Outsource experiment