Purpose To investigate the correlation of the expression of Cyclin D1 and BRAF ( V600E) mutation in papillary thyroid carcinoma ( PTC) and their clinical significance, and to analyze their role in the pathogenesis of PTC. Methods The expression of CK34βE12 and MC protein was detected by immunohistochemical EnVision method in 52 cases of PTC and 52 cases of thyroid benign lesions (25 cases of nodular goiters, 15 cases of Hashimoto’s thyroiditis, 12 cases of follicular adenoma), and the mutation status of BRAF gene protein in above specimens were detected by polymerase chain reaction and DNA sequencing. Expression of Cyclin D1 and BRAF (V600E) mutation in papillary thyroid carcinoma and their correlation clinicopathologic features were also analyzed. Results The positive rate of Cyclin D1 in 88 cases of PTC was 84. 6%,and the immunoreactivity score (4. 6 ± 2. 4), compared with thyroid be-nign lesions (1. 3 ± 1. 6), was statistically significant difference (t=8. 525, P<0. 01). Expression of Cyclin D1 was closely related with lymph node metastasis, capsular invasion, tumor stage Ⅲ+Ⅳ and BRAF (V600E) mutation (P<0. 05), but not related with age, gender, number and diameter of tumor nodules. In 52 cases of PTC BRAF gene mutation rate was 63. 5%, but in thyroid benign lesions mutation rate was 0, with significant difference (P<0. 01). The BRAF mutation is closely related with lymph node metastasis, capsular invasion and tumor stage Ⅲ+Ⅳ(P<0. 05), but not related with age, gender, number and diameter of tumor nodules. The expression of Cyclin D1 in BRAF (V600E) mutation group was stronger, the positive expression rate was significantly higher than that of wild group (100% vs 57. 9%), and expression of the mean score was higher than that of the wild group (5. 7 ± 1. 6 vs 4. 0 ± 2. 5);the comparison between the two groups showed statistical difference (t=2. 652, P<0. 05). Conclusions The expression of Cyclin D1 and BRAF (V600E) mutation are positively related, which are also closely related with the occurrence and development of PTC, and can be used as an index to predict the invasion and metastasis in PTC.

Objective To develop a one-step PCR assay for rapid discrimination of six Brucella species and some intraspecific biovars.Methods Using 6 pairs of primers in one-step PCR to differentiate six classical Brucella species and some biovar in ordinary PCR instrument.The tested strains including 27 reference strains of six Brucella species and 239 Brucella strains were estimated by the PCR assay and biological identification methods.Results The six Brucella species could be precisely differentiated by the onestep PCR assay from the tested strains.Five biovars and vaccine strain of B.suis species could be determined,and biovars 1,3,4 and biovars 2,5,6,7,9 of B.abortus species could be identified at the level of their biovar,moreover,biovars 1,2 and 3,and vaccine strain Rev 1 of B.melitensis species were also discriminated at the biovar and strain level.The accurate rates of the biological identification method and the PCR assay were 98.33％ and 100％ respectively.Conclusion One-step PCR assay was a rapid,specific,and low cost method for identification of Brucella species and discriminating biovars in ordinary PCR instrument.

Objective To establish a RP- HPLC method for quantitative determination of puerarin in Laige Granules. Methods The separation was performed on Econosphere C18 column (4.6 mm? 250 mm,5 ? m) with mobile phase of methanol and 1 % acetic acid solution (25 ∶ 75) at a flow rate of 0.8 mL/min. The UV detection wavelength was 250 nm and the column temperature was 40 ℃ . Results The linear range of puerarin was 0.2～ 1.0 ? g, r = 0.999 7. The mean recovery was 100.56 % (RSD=1.21 % , n = 5 ). Conclusion This method is simple, accurate, sensitive and with good reproducibility .

We constructed prokaryotic recombinant expression vector of P61 gene from Nocardia brasiliensis,expressing P61 protein with biological activity in E.coli,and lay a foundation for further studies related to P61.P61 gene was synthesized and cloned into an expression vector pET-30a(+).The recombinant vector was transformed into Escherichia coli BL21 and induced with IPTG.The production was analyzed with Western blot and the catalase activity of P61 was tested with Catalase Assay Kit.The protein of P61was successfully expressed in E.coli with solubility and high catalase activity,and could be identified by anti-N.brasiliensis sera from mice.The prokaryotic expression plasmid of protein P61 was constructed successfully and can be expressed efficiently in E.coli BL21 cells with higher catalase.

Strain-resource engineering is often considered as an important infrastructure of microbiology related research and industry. The western developed countries took the lead in establishing the classical microbial resource utilization method, and continuously improved the preservation system, species annotation technology and global sharing mechanism, which realized the expansion and reserve of biological resources since end of the 19th century. The rich and diversified germplasm resources, standard strains and production strains not only have important economic values, but also maintain the advantages of scientific research, bioeconomy (such as antimicrobial agents, vaccines, detection reagent development and standard development, etc.) and national security. Although there has been a lot of progress in related research in recent years, compared with developed countries, there is still a big gap in related fields in China. The investment and top-level design in this area lag far behind the western developed countries, and it is not commensurate with the current level of economic and social development in my country. Drawing lessons from the practice of WFCC and WDCM (World Data Center for Microorganisms, Global microbial data Center, affiliated to WFCC), for the purpose of collecting new clinical species/strains, this paper puts forward some suggestions on the identification, preservation and upload system of isolates.