The irradiation of ceils combined with the immunoconjugate of gold nanoparticles by the short pulse laser can make the plasma membrane be transiently permeabilized, which can be used to transfer exogenous molecules into the cells. We explore this technique as a novel gene transfection method for floating cells. Three different floating cells exposed to the laser are selectively transfected with fluorescein isothiocyanatedextran, antibody, and green fluorescent protein （GFP） coding plasmids, and the viability of cells are determined by propidium iodide. For fluorescein isothiocyanate-dextran, the best transfection efficiency of 65% is obtained; for the antibody, it is 74%; whereas for the green fluorescent protein coding plasmids, a very small transfection efficiency is gained. If the transfection efficiency is improved, gold nanoparticles will be very useful as mediator for gene transfection in living cells.
Objective To explore the optimal primer ratio and concentration of asymmetric polymerase chain reaction (A-PCR) in producing hepatitis B virus (HBV) single-stranded DNA (ssDNA) for pyrosequencing. Methods A-PCR was carried out to generate HBV ssDNA with forward to reverse primers of different ratios (50∶1, 100∶1) and concentrations (13.0 pmol/25μL and 0.14 pmol/25μL, 19.5 pmol/25μL and 0.21 pmol/25μL), and the product yield and quality were compared respectively. Results The forward to reverse primer ratio of 50∶1 provided better yield and concentration of 19.5 pmol/25μL and 0.21 pmol//25μL generated a clearer band. Conclusion A simple and feasible method to produce HBV ssDNA for pyrosequencing in batch is established.
Nian-cai Peng, Chun-lin Wang, Li-li Zhang, Mao-lin Lu, Zhen-xi Zhang Institute of Biomedical Analytical Technology and Instrumentation, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an 710049, China.
