Electroporation of siRNA into CLL Cells

Electroporation of siRNA into CLL cells was performed using Amaxa Human B-cell Nucleofection Kit (Amaxa, Cologne, Germany). 1x10⁷ PBMCs were mixed with 100 μL of Amaxa B-cell nucleofector solution, and 2 μg of siRNA was nucleofected using program X-005 [29 (link)]. Transfection efficiency, assessed by transfection with 2 μg pMaxGFP plasmid, was 30–60% with cell viability of 50–80% at 24 hours. siRNA oligos were synthesized by Dharmacon (Lafayette, CO). Sense strand against human ASK1: GCUCGUAAUUUAUACACUGUU.

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Paiva C., Godbersen J.C., Soderquist R.S., Rowland T., Kilmarx S., Spurgeon S.E., Brown J.R., Srinivasa S.P, & Danilov A.V. (2015). Cyclin-Dependent Kinase Inhibitor P1446A Induces Apoptosis in a JNK/p38 MAPK-Dependent Manner in Chronic Lymphocytic Leukemia B-Cells. PLoS ONE, 10(11), e0143685.

Publication 2015

Amaxa Human B-cell Nucleofection Kit siRNA oligos

Ask1 B cell Cell viability Cells Electroporation Human Oligos Plasmid Sirna Transfection

Corresponding Organization : Piramal (India)

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Variable analysis

independent variables

Electroporation of siRNA into CLL cells using Amaxa Human B-cell Nucleofection Kit
Transfection of 2 μg of siRNA using program X-005

dependent variables

Transfection efficiency
Cell viability at 24 hours

control variables

1x10^7 PBMCs mixed with 100 μL of Amaxa B-cell nucleofector solution
Transfection with 2 μg pMaxGFP plasmid

controls

Positive control: Transfection with 2 μg pMaxGFP plasmid
Negative control: Not explicitly mentioned

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