I tried transfecting HAP1 cells with electroporation programme by Lonza (X005,Y007,X007 and X001). Our plasmid size is 8.5 kb. Positive control plasmid eGFP is around 3.5 kb and that shows good transfection efficiency around 80 %. However, incase of our plasmid the transfection efficiency is very low just three to four cells. I tried chemical transfection using Lipofectamine 2000, XtremeGene9, Genjet Still the transfection efficiency is very low Just three to four cells. has anyone transfected HAP1 cells with larger size plasmid around 8.5 kb.
hello
Maybe you need to deal with amount of DNA and lipofectamine volume. Generally we use 1 DNA (µg) for 2 lipo (µL) but you can change and use 1:3 or mor. Because lipofectamine "create" drops with DNA so you need to recovert better the DNA and use more lipo. Be careful, more lipo can kill more cells.
If you have always problem, you can try with JetPrime. Its less rude for cells and you dont need to use Opti MEM. Moreover, SVF in opti MEM inactivate transfection by lipofectamine. I dont know how it works with electroporation, but you can try in cell culture maybe its more easier and less stressfull
Hope that can help you
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