We want to inhibit gene expression via shRNA in prostate cancer cell lines and then store them for a while for further experiments (apoptosis, proliferation, qRTPCR etc.). I know that it is difficult to generate and maintain stable shRNA-inhibited cell lines and shRNAs eager to degrade. Storing them for a period of time can fail the whole experiments. But is there a way to solve this problem? Maybe a chemical solution or technique to postpone the process.
you can use lentivirus that contain the shRNA construct to generate stable cell lines that constantly has the gene knocked down. You could purchase ready to use lentivirus from companies (like Santa Cruz Biotech or horizon discovery) that target your gene of interest.
Hello Ege Sevinç
using shRNA for RNA silencing (i.e., knockdown using transient silencing) can only be employed for a very short time period. The silencing effect is lost after 6-8 days. Since this is not a case of knockout where your gene of interest is removed from the DNA, it is not suggested that you store these cells for further experiments. A batch of transiently transfected cells should only be used for 1 experiment. Alternatively, you can prepare a gene knockout (using CRISPR -Cas or homologous recombination or retroviruses), selected such cells under antibiotic pressure and store them for longer periods to undertake any experimentation.
Hope this answers your question.
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