How to transduce cells to have One-gene-One-cell transduction?

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Weber Lin

Hi Suipwksiow,

For transducing genes into cell by virus, you can adjust the ratio of target cell's number and virus titer (1:0.1~1:0.3) to reach the goal of one gene on one cell. However, I think it is hard to be controlled for one plasmid/gene on one cell by transient transfection.

Best,

Govinda Sharma

Do the ORF cassettes have GFP tag or some sort of other marker?

I usually take a small aliquot of the virus to functionally titer over the target cell line that you'll be working with. Usually this involves infecting and then measuring the % of fluorescent cells you get after 72h.

Once you get a functional titer in TU/mL, I do the large scale library transduction with an MOI of about 0.1 (0.1 TU/cell). This will yield you only about 10% of your cells being transduced but, based on Poisson statistics, >95% of those positive transductants will be singletons.