EGCG联合不同剂量放疗对人肝癌细胞系HepG2中hTERT表达的影响

目的:研究相同剂量表没食子儿茶素没食子酸酯(EGCG)联合不同剂量放疗对HepG2中hTERT基因表达的影响,探讨EGCG成为放疗增敏剂的作用.方法:用MTT比色法检测EGCG对HepG2细胞增殖的影响,用荧光显微镜检测EGCG诱导HepG2的细胞凋亡,从而找到合适的EGCG(25μmol/L)剂量用于观察其放射效果的实验;采用实时荧光定量PCR检测不同剂量放疗(0、2、4、6Gy)联合EGCG和不联合EGCG治疗后,HepG2中hTERT基因的表达.放疗用直线加速器6MVX线.结果:EGCG具有抑制HepG2细胞增殖并诱导HepG2细胞凋亡的作用.同时,不同剂量放疗联合EGCG和不联合EGCG治疗后HepG2中hTERT基因的表达比较,空白组无显著差异;2Gy组、4Gy组、6Gy组均有显著差异(hTERT基因表达的扩增倍数:0.477±0.025vs0.973±0.024;1.110±0.083vs1.382±0.051;1.174±0.128vs1.452±0.109,P<0.01或0.05),并以2Gy组最为显著.结论:EGCG具有抑制HepG2细胞增殖并诱导其凋亡的作用;EGCG联合不同剂量放疗可能下调HepG2中hTERT基因水平,从而抑制端粒酶活性,以放射剂量为2Gy时显著,因此,EGCG有可能成为放疗增敏剂.

Cloning and Expression of Conserved Sequences of cry Gene in E.coli Strain Rosetta(DE3)

[Objective] The aim was to clone the conserved sequences of cry gene and express them in Rosetta （DE3）. [Method] Specific primers were designed according to NCBI database information and the conserved sequences of cry gene were amplified by PCR from Bt transgenic cotton. Then recombinant plasmids were constructed and expressed in E. coil strain Rosetta （DE3）. Finally, the effects of different concentrations and inducing time of IPTG on the expression level of protein were investigated. [Result] Two conserved sequences （304 and 853 bp respectively） of cry gene were amplified. The result of SDS-PAGE confirmed that the recombinant plasmids pGEX-4t-I-304 and pGEX-4t-1-853 could express fusion proteins by IPTG induction and the molecular weight of protein products was 39 and 62.4 kDa respectively, which was in accordance with predicted result. The optimal protein ex- pression conditions were confirmed as induction with 0.15 mmol/L IPTG for 7 h. [Conclusion] This study prepared the ground for the further detection of Bt transgenic crops.

cry基因保守序列克隆及其在大肠杆菌Rosetta(DE3)中的表达

[目的]对cry基因保守序列进行克隆,并使其在大肠杆菌Rosetta(DE3)中进行表达。[方法]根据NCBI数据库信息设计引物序列,采用PCR技术从抗虫棉基因组DNA中扩增出抗虫基因cry的保守序列,构建重组载体并转化到大肠杆菌Rosetta(DE3)菌株中,利用pGEX原核表达系统诱导蛋白表达。同时,分析了不同IPTG浓度及不同诱导时间对蛋白表达量的影响。[结果]扩增出抗虫基因cry的两段保守序列,长度分别为304和853 bp;SDS-PAGE检测结果显示,经IPTG诱导后重组质粒pGEX-4t-1-304和pGEX-4t-1-853成功地表达了大量GST融合蛋白,分子量分别为39和62.4 kDa,与预期结果一致;确定了IPTG最佳诱导浓度为0.15 mmol/L,最佳诱导时间为7h。[结论]为今后检测转Bt基因农作物奠定了基础。

TNF-α与肥胖相关胰岛素抵抗的研究进展

肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）是一种在肿瘤细胞中发挥细胞毒性作用的肽类物质，曾经被认为在体内可使体重减轻而在体外又可促进脂肪生成。1993年，首次在啮齿类动物的脂肪组织中发现了TNF-α，它在肥胖动物模型中的表达量呈显著增加，并且在肥胖人的脂肪组织中也过量表达，而缺乏TNF-α或其受体的肥胖小鼠胰岛素抵抗（insulin resistance，IR）改善。因而初步确定TNF-α也是由脂肪细胞分泌的一种促炎症因子，不仅发挥介导免疫反应、细胞凋亡和存活、细胞毒性以及其他细胞因子的产生的作用，而且与肥胖相关IR的发生密切关联。现将TNF-α与肥胖相关IR的研究进展综述如下。

正交设计优化南苍术快速繁殖培养基的研究

[目的]保护野生苍术资源,并为南苍术工厂化生产提供技术指导。[方法]以南苍术[Atractylodes lancea(Thunb.)DC]种子为材料得到无菌试管苗,应用正交试验法研究6-苄基腺嘌呤(6-BA)、萘乙酸(NAA)、激动素(KT)及其配比对南苍术无菌苗增殖的影响;同时研究了NAA浓度对南苍术苗生根的影响;以泥土、河沙、棉籽壳为材料,采用正交试验法设计9种轻型基质配方,研究其对南苍术生长的影响。[结果]适合南苍术试管苗增殖的培养基为MS+6-BA 1.0 mg/L+NAA 0.4 mg/L+KT 0.4 mg/L;适合南苍术生根的培养基为1/2MS+NAA 0.5 mg/L;最适基质配比为泥土∶河沙∶棉籽壳=3∶3∶2(体积比)。[结论]筛选出了南苍术增殖培养和生根的最佳配方,以及基质的最佳配比,为苍术资源的实验室保存和种苗工厂化生产提供了技术依据。

Study on Rapid Propagation Medium of Atractylodes Lances by Orthogonal Design

[Objective] The aim was to protect wild resource of Atractylodes lances and to provide technical guidance for industrialization of Atractylodes lances. [Method] The test-tube seedlings were produced from seeds of Atractylodis Lances. At the same time, the effects of 6-BA, NAA and KT on the proliferation of aseptic seedling was studied through the orthogonal test. Effect of NAA concentration on the rooting of aseptic seedling was studied through nine types of light substrate groups, which consist of peat soil, fine sand and cotton seed hull, and their effects on Atractylodes lances growth were studied on with orthogonal design. [Result] The culture medium suitable for the proliferation of the Atractylodes lances tube seedling was MS＋6-BA 1.0 mg/L＋NAA 0.4 mg/L＋KT 0,4 mg/L. And the medium suitable for rooting was 1/2MS＋NAA 0.5 mg/L. The optimal ratio of soil, riversand and cotton seed hull was 3：3：2. [Conclusion] The research selected optimum culture medium, rooting medium and ratio of substrate, which provided technical guidance for laboratorial preservation of germplasm resource of Atractylodes lances and industrialization of its seedling production.