Hello!
I am actually planning and siRNA knockdown experiment and therefore im trying to design an siRNA cassette, which expresses for example 6 different siRNAs after transfection with in expression vector in cells.
The design of the cassette im imagening would look like this:
U6-siRNA1-Tstretch-U6-siRNA2-Tstretch [...]
Now my question. Should i add a spacer between Tstretch terminator of siRNA1 and next U6 promotor for next siRNA or should i just construct the cassette that directly after one Tstretch the next U6 sequence starts?
Thank you!
Warm regards, Dominik
Depends how long your siRNA segments are since having the U6 promoters too close together will inhibit transcription.
Robert Adolf Brinzer My siRNA segments will be 21nts sense, 6bp loop and 21nt antisense , T- stretch for terminantion will be 6 T long. So in sum 54bp.
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