siRNA / miRNA gene silencing Human Melanoma cells (501 Mel and SK Mel 28)

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 JetPrime to perform DNA transfection Mammalian cells - Immortalized cell lines HeLa

Get tips on using JetPrime to perform DNA transfection Mammalian cells - Immortalized cell lines COS7

Get tips on using ATP Assay to perform Cell cytotoxicity / Proliferation assay cell type - adipose stem cells

Get tips on using pDG101 to perform Protein Expression Prokaryotic cells - E. coli mScarlet-I

Get tips on using pZH713 to perform Protein Expression Prokaryotic cells - E. coli PP7cp-SYFP2

Get tips on using pFR2 to perform Protein Expression Prokaryotic cells - E. coli ferulic acid

Get tips on using pHYαCGT7 to perform Protein Expression Prokaryotic cells - B. subtilis α-CGTase

Get tips on using pHYαCGT1 to perform Protein Expression Prokaryotic cells - B. subtilis β-CGTase

Get tips on using pAF3 to perform Protein Expression Prokaryotic cells - B. subtilis PHY US417