What are the requirements for direct Nucleofection™ of mRNA for protein expression?


The mRNA should be capped and polyadenylated. The conditions of Nucleofection™ will be the same as for DNA with the particular cell type, i.e. follow the same protocol and use the same program, except one will likely need to add a much higher amount of nucleic acid, 10-20 ug. This must be done in a volume less than or equal to 1/10 of the sample size (so less than 10 µl for 100 µl reaction), so concentration of the mRNA may be necessary. Here is a reference that maybe helpful as well: Nucleofection™ of mRNA (Miyahara et al., Clin Cancer Res (2005) 11(15): 5581-5589; Van De Parre et al., Biochem Biophys Res Commun (2005) 327(1): 356-360; Coughlin et al.; Blood (2004) 103: 2046)