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SEB基因定位突变体构建及其重组表达产物生物学活性研究
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作者 范芳华 严杰 +1 位作者 毛亚飞 于小妹 《浙江医学》 CAS 2010年第12期1741-1746,共6页
目的 构建葡萄球菌肠毒素B(SEB)基因及其突变体原核表达系统,了解突变前、后重组表达产物的细胞毒性、促淋巴细胞增殖和抑制肿瘤生长活性的变化.方法 采用突变引物PCR构建SEB基因定位突变体,建立SEB基因及其定位突变体原核表达系统;... 目的 构建葡萄球菌肠毒素B(SEB)基因及其突变体原核表达系统,了解突变前、后重组表达产物的细胞毒性、促淋巴细胞增殖和抑制肿瘤生长活性的变化.方法 采用突变引物PCR构建SEB基因定位突变体,建立SEB基因及其定位突变体原核表达系统;同时采用Ni-NTA亲和层析法提纯表达的目的 重组蛋白;采用TCID50法测定目的 重组蛋白对Vero细胞的毒性;采用MTT比色法分别检测不同浓度的目的 重组蛋白促使小鼠脾细胞增殖及对抑制KB和HL-60癌细胞株生长的作用.结果 所克隆的SEB基因核苷酸序列与各项研究报道的相似性为100%,3种突变体均在既定位置获得预期的密码子突变.rSEB和各突变体重组蛋白表达量约为细菌总蛋白的40%;rSEB对Vero细胞TCIC50为3.4μg,其突变体重组蛋白分别为3.2~16.8μg;1和5μg/ml的rSEB及突变体重组蛋白rSEB/D9N对小鼠脾细胞均有明显的促增殖作用(P<0.05),10和20μg/ml的rSEB及rSEB/D9N促进小鼠脾细胞增殖活性与100μg/ml植物血凝素PHA相似(P>0.05).5~20μg/ml的rSEB及rSEB/D9N作用的小鼠脾细胞上清液及其上清液与脾细胞混合物均能有效抑制KB和HL-60细胞生长(P<0.05).结论 本研究成功构建了SEB基因突变体及其高效原核表达系统;其中突变体重组蛋白rSEB/D9N细胞毒性较低,促脾细胞增殖和抑制肿瘤细胞生长活性较强,可作为研制升高白细胞、抗肿瘤的SEB相关药物的候选突变体. 展开更多
关键词 葡萄球菌肠毒素B SEB基因/定位突变原核表达系统/构建重组蛋白/细胞毒性
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Localization of Two GFP_tagged Tobacco Plastid Division Protein NtFtsZs in Escherichia coli 被引量:1
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作者 王东 孔冬冬 +3 位作者 鞠传丽 胡勇 何奕昆 孙敬三 《Acta Botanica Sinica》 CSCD 2002年第8期931-935,共5页
Two plastid division genes, NtFtsZ1 and NtFtsZ2 isolated from Nicotiana tabacum L. were fused with gfp and expressed in Escherichia coli . The regular localizations of full length NtFtsZs∶GFP along the fil... Two plastid division genes, NtFtsZ1 and NtFtsZ2 isolated from Nicotiana tabacum L. were fused with gfp and expressed in Escherichia coli . The regular localizations of full length NtFtsZs∶GFP along the filamentous bacteria indicated that the NtFtsZs could recognize the potential division sites in E. coli and be polymerized with heterogeneous FtsZ from bacteria. The overexpression of NtFtsZs ∶ gfp inhibited the division of host strain cells and resulted in the long filamentous bacterial morphology. These results suggested that eukaryotic ftsZs have similar function to their prokaryotic homologs. Meanwhile, the different deletions of motifs of NtFtsZs are also employed to investigate the functions of these proteins in E. coli . The results showed that the C_terminal domains of NtFtsZs were related to the correct localization of NtFtsZs in E. coli and the N_terminal domains of NtFtsZs were responsible for the polymerization of homogeneous and heterogeneous FtsZ proteins. The significance of these results in understanding the functions of NtFtsZs in plastid division were discussed. 展开更多
关键词 Nicotiana tabacum plastid division gene NtFtsZ GFP localization in Escherichia coli
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Cloning and Expression Analysis of the High Mobility B Group Genes in Arabidopsis thaliana 被引量:1
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作者 冀芦沙 王洪霞 郭尚敬 《Agricultural Science & Technology》 CAS 2010年第8期160-163,172,共5页
[Objective] The aim was to better research the function and action mode of High Mobility Group B (HMGB) proteins in higher plants. [Method] At2G33450,At5G23405 and At5G23420 genes of HMGB protein family in Arabidops... [Objective] The aim was to better research the function and action mode of High Mobility Group B (HMGB) proteins in higher plants. [Method] At2G33450,At5G23405 and At5G23420 genes of HMGB protein family in Arabidopsis thaliana were cloned by the use of RT-PCR method,and the expression of these three proteins in E.coli and Arabidopsis thaliana were detected by using SDS-PAGE,Northern blot and subcellular localization methods. [Result] The results showed that the molecular weights of the three proteins were 17.5,17.0 and 27.0 kD respectively,and the expression levels of the proteins in Arabidopsis thaliana were At5G23420At5G23405At2G33450. In addition,all the three proteins were located in nucleus. [Conclusion] The study will provide a basis for the further research on the biological function of HMGB proteins in higher plants. 展开更多
关键词 Arabidopsis thaliana High Mobility Group B (AtHMGB) Prokaryotic expression Subcellular localization Northern blot
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Synthesis, in-situ coating and characterization of scorodite with high leaching stability 被引量:3
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作者 Ping-chao KE Zhi-hong LIU 《Transactions of Nonferrous Metals Society of China》 SCIE EI CAS CSCD 2019年第4期876-892,共17页
To improve stability of scorodite,a method of simultaneous synthesis and in-situ coating of scorodite was proposed.Scorodite particles with polyhedral and raspberry-like morphologies were synthesized in an Fe(Ⅱ).As(... To improve stability of scorodite,a method of simultaneous synthesis and in-situ coating of scorodite was proposed.Scorodite particles with polyhedral and raspberry-like morphologies were synthesized in an Fe(Ⅱ).As(Ⅴ).H2O system at 90℃and pH 1.5 by blowing oxygen gas into the system.When the initial Fe/As molar ratio exceeded 1:1,a coating of sulfate-containing iron(hydr)oxides formed on the surfaces of scorodite particles during synthesis.To evaluate the leaching stability of synthesized scorodite samples,toxicity characteristic leaching procedure(TCLP)tests were conducted at pH 4.93 for 60 h,and long-term leaching tests were conducted for 30.40 d within a pH range of 5.40.10.88.The leaching results indicated that the release of arsenic from scorodite was noticeably postponed by the coating,and the average arsenic concentrations in the leaching solutions were as low as 0.12 mg/L in the TCLP tests and lower than 0.5 mg/L in the long-term leaching tests. 展开更多
关键词 SCORODITE arsenic immobilization NUCLEATION in-situ coating STABILITY
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Chromosomal localization of a tandemly repeated DNA sequence in Trifolium repens L
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作者 ZHU JM NW ELLISON +1 位作者 RE ROWLAND (The Horticulture and Food Research Institute of NewZealand, M Albert Research Centre, Private Bag 92169, Auckland, New Zealand.)(New Zcaland Pastoral Agriculture Research Institute, Grasslands Research Centre, Private Bag 11 《Cell Research》 SCIE CAS CSCD 1996年第1期39-46,共8页
A karyotype of Trifolium repens constructed from mitotic cells revealed 13 pairs of metacentric and 3 pairs of submetacentric chromosomes including a pair of satellites located at the end of the short arm of chromosom... A karyotype of Trifolium repens constructed from mitotic cells revealed 13 pairs of metacentric and 3 pairs of submetacentric chromosomes including a pair of satellites located at the end of the short arm of chromosome 16.C-bands were identified around the centromeric regions of 8 pairs of chromosomes. A 350 bp tandemly repeated DNA sequence from T. repens labelled with digoxygenin hybridized to the proximal centromeric regions of 12 chromosome pairs. Some correlation between the distribution of the repeat sequence and the distribution of C-banding was demonstrated. 展开更多
关键词 Tandem repeat in situ hybridization digoxygenin-labelled probe C-BANDING KARYOTYPE
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CELLULAR LOCALIZATION AND EXPRESSION OF pygo DURING DROSOPHILA DEVELOPMENT
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作者 LINXin-da LINXin-hua CHENGJia-an 《Entomologia Sinica》 CSCD 2003年第2期95-102,共8页
Wg/Wnt signaling is a key signaling pathway in Drosophila. Many genes involved in Wingless(wg) signal transduction pathway downstream of Wg, or it'' s vertebrate Wg homologue Wnt, have been identified.Transduc... Wg/Wnt signaling is a key signaling pathway in Drosophila. Many genes involved in Wingless(wg) signal transduction pathway downstream of Wg, or it'' s vertebrate Wg homologue Wnt, have been identified.Transduction of the Wg signal downstream of Wg is mediated by nuclear TCF/LEF-1, through association with Ar-madillo (Arm)/β-catenin. Pygopus (pygo) is a new identified component in this pathway . Cellular localization experiment showed that pygo was expressed specifically in the nucleus. The expression profile of pygo in embryos was examined using in situ hybridization. Although pygo expressed ubiquitously in the embryos, it expressed at relatively high level in pre-blastoderm embryos which indicate a high degree of maternally provided message, fol-lowed by a low level of ubiquitous zygotic expression. This continues into larval tissues (including wing disc, eye disc and leg disc), where pygo appears to be expressed at low level. Comparison of pygo expression levels, in the wing disc, eye disc and leg disc, showed pygo expression level in the wing disc pouch and leg disc were rela-tive higher. 展开更多
关键词 DROSOPHILA WINGLESS WNT Pygo NLS in situ hybridization
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