Efficient DNA transfection in neuronal and astrocytic cell lines.
We have studied different parameters for efficient DNA transfection in various cell types and with different size of the promoter. Here we report that the optimum condition for DNA transfection by electroporation is 350 V/960 microF for PC12, 450V/960 microF C6 cells, and 250 V/500 microF for COS-1 cells. For the human neuroblastoma (SK-N-SH) cells the optimum condition for DNA transfection is by the calcium phosphate method. In promoter mapping studies, a serial deletion approach is commonly used. To optimize transfection we have selected three DNA constructs that varied in size from 4.5 to 12.4 kilobases (kb). We measured the promoter activity of these constructs under conditions of 'equal amount', 'equimolar', and 'equimolar plus carrier DNA to make it equal amount'. We recommend that for comparative purpose, transfection should be carried out under 'equimolar condition' without a need to adjust the total amount of DNA by carrier DNA. Taken together, our results suggest that efficient methods for DNA transfection are important to study gene regulation by devising better ways to deliver DNA into the mammalian cells.
Ghosh
C
C
Institute of Psychiatric Research, Department of Psychiatry, Indiana University School of Medicine, Indianapolis 46202, USA.
Song
W
W
Lahiri
D K
DK
D016564:Amyloid beta-Protein Precursor / Q000235:genetics; D000818:Animals; D001254:Astrocytoma / Q000235:genetics; D019556:COS Cells; D002460:Cell Line; D016543:Central Nervous System Neoplasms / Q000235:genetics; D004247:DNA / Q000737:chemistry; D004285:Dogs; D018274:Electroporation / Q000379:methods; D006367:HeLa Cells; D006801:Humans; D009474:Neurons / Q000166:cytology* / Q000502:physiology*; D016716:PC12 Cells; D051381:Rats; D014162:Transfection / Q000379:methods*
D016564:Amyloid beta-Protein Precursor; D004247:DNA