Protein Expression Eukaryotic cells Hi5

Get tips on using Gibco™ IMDM, GlutaMAX™ Supplement to perform 3D Cell Culture Media hiPSC-derived lung organoids

Hello! I used Trizol to extract total RNA from in-vitro cultured bacteria (1 X 10^8 cells). After phase separation, I mixed ~0.4 ml of the upper phase which contains RNA with 0.5 mL cold isopropanol. However, the amount of RNA when measured in Nanodrop was very low. In addition, the ratio between 260 and 230 was around 0.1 to 0.5. Is there a chance that my sample was contaminated by the Trizol reagent? When I collected the aqueous phase I made sure to not touch the lower phase. What should I do?

Get tips on using Gibco™DMEM/F-12, no glutamine to perform 3D Cell Culture Media hiPSC-derived lung organoids

Get tips on using Gibco™KnockOut™ DMEM to perform Stem cell Differentiation media hiPSCs or hESCs differentiation to Embryoid body (EB)

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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, no glucose, no glutamine, no phenol red to perform 3D Cell Culture Media hiPSC-derived cardiac organoids

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Get tips on using Corning® 500 mL Iscove’s Modification of DMEM to perform 3D Cell Culture Media hiPSC-derived lung organoids

Get tips on using Gibco™ StemPro™ hESC SFM to perform Stem cell Differentiation media hiPSCs or hESCs differentiation to Embryoid body (EB)