Depu Chen

DNA purification and gene typing: Based on multifunctional nanobeads

In this report, a universal protocol for extracting genomic DNA from whole blood, saliva, and bacterial culture by using magnetic nanobeads as solid-phase absorbents was presented. The enrichment of target cells and adsorption of DNA have been functionally integrated onto the surfaces of the carboxyl-modified magnetic nano-beads, and the DNA segments bound on the surface of the beads can be directly used as PCR templates to amplify a target gene. The PCR products were applied to an oligonucleotide array to perform gene typing. The protocol proves to be simple, rapid, biologically and chemically nonhazardous, and promising for the microfabrication of DNA preparation chip.

Assaying poly(ADP-ribose) polymerase activity in plants by polarographic method

A new method has been developed to assay poly(ADP-ribose) polymerase (PARP) activity in plant tissues through determining the content of nicotinamide (NIC) produced by enzymatic reaction by linear sweeping polarographic method. The detection limit of NIC was 0.03 μmol/L, the calibration graph was linear up to 5 μmol/L (r = 0.999). The recoveries were approximately in the range of 92% to 98% and the relative standard deviations were less than 6.6%. Moreover, NAD+ and other interference existing in the mixture after enzymatic reaction had been removed by simple pretreatment, thus PARP assays were not interfered. A rapid, simple, sensitive and reliable nonisotopic method is reported to assay PARP activity in plant tissues. The results show that the KmNAD value of PARP in maize (Zea mays L.) seedlings is 59 and the optimum pH for PARP activity is 8.5. Moreover, physiological conditions affect PARP activity in plant tissues. which has not been reported previously. When tobacco (Nicotiana tobacum) suspension cells were stressed by NaCl at low concentrations (100, 200 mmol/ L), the PARP activity increased significantly; when the cells were stressed at high concentrations (400, 1 000 mmol/L), it decreased to or even below the control level. PARP activity in etiolated maize seedlings was higher than that in light-grown seedlings.