内皮抑素-血管内皮细胞抑制因子重组腺病毒对新生血管的抑制效应

内皮抑素与血管内皮细胞抑制因子均为内源性新生血管抑制分子。为了探讨两分子融合后的生物学功能,以重组腺病毒为载体介导融合基因在体内外表达。体外实验表明重组腺病毒Ad-hENDO-VEGI151能够在ECV304、HepG2、L929等细胞中表达41000大小的融合蛋白,对血管内皮细胞增殖有明显的特异性抑制作用,对HepG2和L929细胞无抑制作用(F=13112.13,P=0.0001)。体内实验表明Ad-hENDO-VEGI151重组腺病毒表达的融合蛋白可以显著抑制鸡胚绒毛尿囊膜血管的生成;与AdLacZ对照组相比,Ad-hENDO-VEGI151对兔炎症性角膜新生血管的抑制明显(F=1413.11P=0.0001),免疫组化结果显示,融合蛋白主要在角膜上皮层表达;治疗荷肝癌裸鼠,注射Ad-hENDO-VEGI151的肿瘤体积(487.7±241.2mm3)与AdLacZ对照组(4075.9±1849.9mm3)差异显著(F=14.80P=0.0085),抑瘤率达到88.03%,免疫组化结果显示,胞膜染色阳性,Ad-hENDO-VEGI151组肿瘤的微血管密度30.75±3.31%与AdLacZ组50.25±8.65%差异明显F=17.72P=0.0056抑制率为39%。结果提示:重组腺病毒Ad-hENDO-VEGI151能够表达有生物学活性的融合蛋白,并发挥特异性的新生血管抑制作用。     

小分子蛋白骨架提高噬菌体展示肽亲和力

本综述小分子蛋白骨架在提高哓菌体展示肽亲和力方面的作用。目前已知的天然及人工合成的小分子蛋白骨架包括α-螺旋结构,β-折叠片复合结构,α-螺旋结构展示肽亲和力高达87pmol/L,比野生型高12500倍,β-折叠片结构展示肽亲和力也达到20pmol/L,比野生型高600倍,α-螺旋与β-折叠片复合结构主要是锌指结构域,具有结构复杂,亲和力高的特点,小分子蛋白骨架主要通过提高展示肽的空间构象和对蛋白酶的稳定性而提高亲和力,展现出良好的应用前景。     

L-型细菌蛋白表达系统

L-型细胞由于缺乏细胞壁以及具有的其它特殊生物学特性,通过连接合适的信号肽,可以用于许多外源蛋白的可溶性,功能活性形式的分泌表达,表达产物在培养基中,表达的产量依赖于不同的基因序列,载体,宿主和诱导表达条件等因素,此外,L-型细菌还能将具有功能活性的蛋白展示在胞浆膜表面。     

根癌土壤杆菌介导的丝状真菌转化*

最近研究表明,Agrobacteriumtumefaciens介导转化(ATMT)的方法,可以应用到丝状真菌中。本文将从ATMT的转化原理、转化特点、转化方法以及其在丝状真菌中的主要转化实例四个方面来着重介绍A.tumefaciens介导的丝状真菌转化的最新研究进展。并对其今后的应用前景提出了展望。     

碱性Krueppel样因子对和珠蛋白基因的转录调控作用

为探索碱性Kruppel样因子（basic Krueppel-like factor,BKLF)对γ-和ε珠蛋白基因的转录调控作用,构建了人类BKLF(hBKLF)基因全长及N端缺失40个氨基酸的真核表达载体,并将其瞬时转染到COS7细胞中做报告实验。结果表明,hBKLF能够抑制γ-和ε-珠蛋白基因启动子的活性,而hBKLF特异的反义核酸则激活这两种基因的启动子。N端缺失40个氨基酸不影响这种转录抑制功能。hBKLF强烈抑制FKLF(embryonic/fetalβ-like globin gene-activating Krueppel-like factor)引起的对γ-和ε-珠蛋白基因的转录激活作用。另外BKLF基因能够与SHP1（SH2-containing protein tyrosine phosphatase1)基因启动子区的CACCC元件结合。这些结果提示γ-和ε-珠蛋白基因可能是BKLF的转录调控对象,为研究BKLF参与血细胞特异表达基因的调控提供了依据。     

卵母细胞特异表达φC31整合酶载体pZP3-INT的构建及其在小鼠卵母细胞中的表达

链霉菌噬菌体φC31整合酶是一种位点特异性重组酶(Site-specific recombinase,SSR),可介导链霉菌噬菌体attP位点(Phage attachment site)和链霉菌基因组attB位点(Bacterial attachment site)间的单向重组.为探讨它能否应用于卵母细胞特定基因的重组,文章采用卵巢针刺取卵法采集生发泡(GV)期小鼠卵母细胞,将卵透明带糖蛋白3(ZP3)启动子驱动φC31整合酶表达载体pZP3-INT和检测φC31整合酶位点特异性重组功能的重组质粒载体pBCPB+,通过显微注射导入到小鼠卵母细胞中.培养48 h后,RT-PCR检测φC31整合酶mRNA表达以及PCR检测pBCPB+载体发生重组的情况.结果表明:载体pzP3-INT在卵母细胞中表达φC31整合酶mRNA;并且pBCPB+载体发生了位点特异性重组,提示φC31整合酶在卵母细胞中可以介导位点特异性重组反应.     

Construction and Application of Efficient Ac-Ds Transposon Tagging Vectors in Rice

Transposons are effective mutagens alternative to T-DNA for the generation of insertional mutants in many plant species including those whose transformation is inefficient. The current strategies of transposon tagging are usually slow and labor-intensive and yield low frequency of tagged lines. We have constructed a series of transposon tagging vectors based on three approaches： （i） AcTPase controlled by glucocorticoid binding domain/VP16 acidic activation domain/Gal4 DNA-binding domain （GVG） chemical-inducible expression system; （ii） deletion of AcTPase via Cre-lox site-specific recombination that was initially triggered by Ds excision; and （iii） suppression of early transposition events in transformed rice callus through a dual-functional hygromycin resistance gene in a novel Ds element （HPT-Ds）, We tested these vectors in transgenic rice and characterized the transposition events. Our results showed that these vectors are useful resources for functional genomics of rice and other crop plants. The vectors are freely available for the community,     

The Role of Viral Infection in Inducing Variability in Virus-Free Progeny in Tomato

The effect of virus-host interactions on subsequent generations is poorly understood. The evaluation of the effects of viral infection on inheritance of quantitative traits in the progeny of infected plants and elucidation of a possible relationship between chiasma frequency in the infected plants and variability of traits in the progeny were investigated. The current study involved genotypes of four intraspecific hybrids of tomato （Solanum lycopersicum L.）, their parental forms and two additional cultivars. Used as infection were the tobacco mosaic virus （TMV） and potato virus X （PVX）. The consequences of the effect of viral infection were evaluated based on chromosome pairing in diakinesis and/or by examining quantitative and qualitative traits in the progeny of the infected tomato plants. Tomato plants infected with TMV ＋ PVX were found to differ in chiasma frequency per pollen mother cell or per bivalent. Deviations have been observed for genotypes of both F1 hybrids and cultivars. At the same time, differences in mean values of the traits under study have only been found for progeny populations （F2-F4） derived from virus-infected F1 hybrids, but not in the case of progeny of the infected cultivars. The rate of recombinants combining traits of both parents increased significantly （2.22-8.24 times） in progeny populations of hybrids infected with TMV＋PVX. The above suggests that the observed effects could be the result of modification of recombination frequencies that can be manifested in heterozygous hybrids and make small contributions to variability in cases of ＇homozygous＇ tomato genotypes （i.e. cultivars）.