Genotyping cytomegalovirus UL97 mutations by high-resolution melting analysis with unlabeled probe.

Human cytomegalovirus (CMV) is an opportunistic pathogen, and infections with this virus can be treated with ganciclovir (GCV). Most GCV-resistant clinical CMV isolates contain a mutation in the UL97 gene. Genotypic assays for diagnostic screening of GCV-resistant CMV have been developed. High-resolution melting analysis (HRMA) with unlabeled probe is considered a perfect tool for this purpose. In this study, we have developed an HRMA-based genotypic test for the detection of UL97 mutations. Wild type and M460V/I mutants of UL97 were constructed. HRMA with unlabeled probe was used as a genotyping method for the detection of M460V/I mutations. The melting peaks obtained directly from PCR products did not enable us to distinguish the wild type from M460 mutants. The sensitivity and accuracy of HRMA were dramatically improved by using unlabeled probe. HRMA with unlabeled probe successfully distinguished M460V from M460I and served well for the detection of M460V/I mutations in clinical samples. HRMA with unlabeled probe proves to be a sensitive and cost-effective genotyping method for the detection of M460 mutations.

[Analysis of reproductive ecological factors of endangered Rhodiola gannanica.] / ç”˜å—çº¢æ™¯å¤©æ¿’å±çš„ç”Ÿæ®–ç”Ÿæ€å­¦å› ç´ åˆ†æž.

To find effective ways for protecting and recovering an endangered species Rhodiola gannanica in Gannan, an important distribution region of this species in the eastern Tibetan Plateau of China, the breeding system, reproductive process, flowering phenological characteristics and other reproductive controlling factors were investigated. The results showed that there was a unique breeding system in R. gannanica, which was dioecious, and male flowers were also bisexual plants in the early stage of flower bud differentiation. While the sac of pistil was aborted in a certain stage with the development of flowers, the bisexual plants only played the role of males' functions. The outcrossing reproduction was observed, the pollination mode was mainly by wind, and the population flowering occurred in the season with abundant precipitation and high temperature. The bud satge appeared in early June, the florescence was in mid-June, and the flourishing florescence was in early July. The flo-wering duration of R. fastigiata was about 36 d, and the fruit ripening began in late August. The flo-wering time and the flowering duration of the bisexual plants were earlier than female plants. Due to the low pollination rate of the female plants, the buds and ovules were damaged and the 1-3 years young plants barely got flowering and fruiting, the rate of flowering and fruiting of the community only reached 11.0% and the rate of seed natural propagation was low (about 2.0%). 20.0%-25.1% buds of female plants were damaged from bud to flowering stages. 51.1%-65.0% flowers were aborted from flowering to fruiting stages. Only 10.1%-21.0% of ovules developed into seeds. The seed production of per female plant was 158.1, seed germination rate was 81.5% under artificial conditions, and seedling survival rate reached 36.0% in the first year. Our results revealed that seed quality was not the key ecological factor, while it was the pollen limitation, low seed production and the survival rate of seedling that resulted in the reproductive success and species endangered situation.

The 8-kDa dynein light chain binds to p53-binding protein 1 and mediates DNA damage-induced p53 nuclear accumulation.

The tumor suppressor protein p53 is known to undergo cytoplasmic dynein-dependent nuclear translocation in response to DNA damage. However, the molecular link between p53 and the minus end-directed microtubule motor dynein complex has not been described. We report here that the 8-kDa light chain (LC8) of dynein binds to p53-binding protein 1 (53BP1). The LC8-binding domain was mapped to a short peptide segment immediately N-terminal to the kinetochore localization region of 53BP1. The LC8-binding domain is completely separated from the p53-binding domain in 53BP1. Therefore, 53BP1 can potentially act as an adaptor to assemble p53 to the dynein complex. Unlike other known LC8-binding proteins, 53BP1 contains two distinct LC8-binding motifs that are arranged in tandem. We further showed that 53BP1 can directly associate with the dynein complex. Disruption of the interaction between LC8 and 53BP1 in vivo prevented DNA damage-induced nuclear accumulation of p53. These data illustrate that LC8 is able to function as a versatile acceptor to link a wide spectrum of molecular cargoes to the dynein motor.