Study on the cDNA Library Construction Method for Xinjiang Artemisia rupestris / 中国药房

OBJECTIVE:To establish a method for full-length cDNA library of Xinjiang Artemisia rupestris. METHODS:Mod-ified Trizol method was adopted to extract total RNA in young leaves of A. rupestris,it was transcribed into single-strand cDNA, and then synthesized into double-strand cDNA by long-distance polymerase chain reaction(LD-PCR)method. PCR product was di-gested by proteinase K and sfiⅠ,and then fractionated by CHROMA SPIN-400 columns. The cDNA longer than 0.4 kb were col-lected and ligated to phage λTriplE × 2,and then protein packaging was performed. Full-length cDNA library was established by SMART technology. 20 monoclonal were randomly selected from the library,and electrophoresis was used to determine the primary library titer,library capacity,recombinant positive rate and length of insert cDNA. RESULTS:The primary library titer was 1.94× 107 pfu/mL,library capacity was 0.97×107 pfu;recombinant positive rate of insert cDNA was 96% and length was 0.5-2.0 kb with an average of 0.9 kb. CONCLUSIONS:The established library is high in capacity and quality,which can provide basis for estab-lishing cDNA library of Xinjiang A. rupestris.

Characterization and immunoprotective effect of SjIrV1, a 66 kDa calcium-binding protein from Schistosoma japonicum / 生物工程学报

Calcium-binding protein is an indispensable protein which performs extensive and important functions in the growth of Schistosoma japonicum. Based on our primary study on tegument surface proteins of S. japonicun, a cDNA encoding a 66 kDa calcium-binding protein of S. japonicum (Chinese strain) was cloned, sequence analysis revealed that it was identical with that of SjIrV1 of Philippines strains S. japonicum. The expression of SjIrV1 were detected by Real-time PCR, using cDNA templates isolated from 7, 14, 21, 28, 35 and 42 days worms and the results revealed that the gene was expressed in all investigated stages, and the mRNA level of SjIrV1 is much higher in 42 d female worms than that in 42 d male worms. The cDNA containing the open reading frame of IrV1 was subcloned into a pET28a (+) vector and transformed into competent Escherichia coli BL21 for expression. The recombinant protein was purified using a Ni-NTA purification system, and confirmed by high performance liquid chromatography (RP-HPLC) and tandem mass spectrometry (MS/MS). Western blotting analysis showed that recombinant SjIrV1 (rSjIrV1) could be recognized by the S. japonicum infected mouse serum and the mouse serum specific to rSjIrV1, respectively. Immunofluorescence observation exhibited that SjIrV1 was mainly distributed on the tegument of the 35-day adult worms. ELISA test revealed that IgG, IgG1 and IgG2a antibodies are significantly increased in the serum of rSjIrV1 vaccinated mice. The study suggested that rSjIrV1 might play an important role in the development of S. japonicum.

Preliminary study on virtual touch tissue quantification for evaluation of small focal liver lesions / 中华超声影像学杂志

Objective To evaluate the application value of virtual touch tissue quantification (VTQ)for evaluating small focal liver lesions.Methods Seventy-seven patients with 89 small focal liver lesions,whose largest diameter were less than or equal to 3 cm were evaluated with VTQ.The shcar wave velocity (SWV) of the tumor and background liver parenchyma were calculated,and their results were compared with those of 40 healthy subjects.Statistical analysis was performed on the SWV for the differentiation of normal liver,background liver parenchyma and lesions.The cut-off point for SWV was obtained by using ROC curve analysis which was used to predict malignancy lesions.Results The SWV of malignant lesions was significantly higher than that of benign ones [(2.56-± 0.67) m/s vs (1.70 ± 0.55) m/s,P =0.000].The SWV of metastatic liver carcinoma which came from lung cancer and breast cancer were higher than that of gastrointestinal tract cancer transfers (P =0.000).Based on 1.93 m/s as the cut-off point of SWV value,the sensitivity,specificity,accuracy,positive predictive value and negative predictive value for diagnosing malignancies were 82.0 ％,84.6％,83.15 ％,87.23 ％ and 78.57 ％ respectively.Conclusions VTQ technique can help the differential diagnosis of small focal liver lesions.