TissueFAxs 53BP1 [H-300] Rabbit

ROS has a very short half-lives in biological environment as they are influenced by exposure to ambient oxygen. As it is highly reactive and hard to measure care should be taken to ensure the stability of the sample during isolation, preparation, storage, and analysis.

ROS has a very short half-lives in biological environment as they are influenced by exposure to ambient oxygen. As it is highly reactive and hard to measure care should be taken to ensure the stability of the sample during isolation, preparation, storage, and analysis.

Short hairpin or small hairpin RNA (shRNA) is artificial RNA, which has a hairpin loop structure, and uses inherent microRNA (miRNA) machinery to silence target gene expression. This is called RNA interference (RNAi). These can be delivered via plasmids or viral/bacterial vectors. Challenges in shRNA-mediated gene silencing include: 1. Off-target silencing, 2. Packaging shRNA encoding lentivirus, and 3. Stable transduction in cells. RNAi have been designed to have anywhere from 19-27 bs, but the most effective design has 19 bp. In case commercial shRNAs are not available, potential target sites can be chosen within exon, 5’- or 3’ UTR, depending on which splice variants of the gene are desired. One should use the latest algorithms and choose at least two different sequences, targeting different regions, in order to have confidence in overcoming off-target effects. A BLAST search after selecting potential design will eliminate potential off-target sequences. For the second challenge, sequencing the vector using primers for either strand (50-100 bp upstream) is suggested, along with using enzymatic digestion on agarose gel for the vector. Next, once the shRNA-containing vector is packaged in a virus, it is important to check the viral titer before transduction. Finally, using a marker in the lentiviral vector (fluorescent protein or antibiotic resistance), along with qPCR for target gene expression can help in determining efficacy of transduction and shRNA on its target site.

Get tips on using MACSprep™ HLA B/T Cell Isolation Kit, human to perform Cell Isolation HLA B/T

Get tips on using ON-TARGETplus Human CYP3A4 (1576) siRNA - SMARTpool to perform siRNA / miRNA gene silencing Human - Primary Human Hepatocytes CYP3A4

Get tips on using ON-TARGETplus Human CYP2B6 (1555) siRNA - SMARTpool to perform siRNA / miRNA gene silencing Human - Primary Human Hepatocytes CYP2B6

Get tips on using Anti-Histone H3 (acetyl K9) antibody - ChIP Grade (ab4441) to perform ChIP Anti-bodies H3K9-Ac

Get tips on using Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade to perform ChIP H3K36Me3 - Sheep Rat -NA-

Get tips on using ON-TARGETplus Human NCR3LG1 (374383) siRNA - SMARTpool to perform siRNA / miRNA gene silencing Human - HT-29 B7-H6/NCR3LG1

Get tips on using Anti-Histone H4 (mono methyl K20) antibody - ChIP Grade (ab9051) to perform ChIP Anti-bodies H4K20me1