DNA methylation profiling Gene specific profiling HEK 293T

Get tips on using EMD Millipore™ Chemicon™ CpGenome™ Universal DNA Modification Kit to perform DNA methylation profiling Gene specific profiling - Hep3B SFRP3

Get tips on using EZ DNA Methylation kit to perform DNA methylation profiling Whole genome profiling - OVCAR-3 human ovarian cancer

Get tips on using EZ DNA Methylation kit to perform DNA methylation profiling Whole genome profiling - mouse T-cell (CD4 / CD8)

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 EpiTect Bisulfite Kit to perform DNA methylation profiling Gene specific profiling - MG-63 miR-34a

Get tips on using EpiTect MSP Kit to perform DNA methylation profiling Gene specific profiling - Human ovarian tissue MEG3

Get tips on using MethylEasy™ Xceed (ME002) to perform DNA methylation profiling Gene specific profiling - UMR-106 BMP2

Get tips on using Global DNA Methylation ELISA to perform DNA methylation profiling Whole genome profiling - HCT116, HTC15 human colon cancer cells

Get tips on using EZ-96 DNA Methylation-Gold™ Kit to perform DNA methylation profiling Whole genome profiling - human placenta

Get tips on using EpiTect Methyl II PCR Assays to perform DNA methylation profiling Gene specific profiling - MCF-7 FOXA2