癫痫基因研究进展

本综述了癫痫基因的定位,癫痫候选基因的筛查和癫痫相关基因的寻找及其进展。     

单基因肥胖的致病基因概述

肥胖是人类体质研究中的重要组成部分,发生率在世界范围内呈快速上升趋势,其预防与控制是当今研究的重要课题。肥胖受遗传与环境因素的共同作用,其中遗传因素对其发生至关重要。截至目前,已有14种单基因肥胖的致病基因被确认,涉及了食欲调控和食物摄取、能量消耗和脂肪细胞分化调控等方面。本文概述了上述基因在单基因肥胖研究中的情况,旨在加深对肥胖病理生理学机制的理解,并为其相关研究的开展提供参考。     

Dicer基因

Dicer编码RNaseⅢ家族,是一进化保守基因。它包含四个ORF框架：DExH/DEAH盒解旋酶,PAZ,两个RNaseⅢ催化区与双链RNA结合区DS－RBD。目前发现Dicer在RNA干扰以及异时发育中起重要作用,它可以切割长的双链RNA成长度为22nt左右的小干扰RNA－siRNA,异时基因stRNA的成熟也与需要Dicer的参与。Dicer行使切割功能需要RDE－1或其同源基因ALG1／ALG2的协同。随着研究的深入,Dicer的其它生物功能将进一步被发现。     

基因重组与基因合成实验设计软件系统的建立

本文报导了用于基因重组与基因合成实验设计的软件系统的建立.此系统由30个功能模块组成,为研究者提供了包括在DNA分子上寻找限制性内切酶位点、核酸分子片段之间同源性比较,基因化学合成的实验设计、特定顺序分析引物及核酸杂交探针的设计、阅读框的查找等功能.此外,本系统可以对外来数据库的资料进行援引和进一步分析,为分子生物学的研究提供有价值的信息.     

Class I patatin基因5′侧翼区驱动的GUS基因在马铃薯块茎中专一性表达

将1.4kb Class I patatin基因的5′侧翼区与GUS基因融合,构建了双元表达载体pPATIs(含patatin部分信号顺序)和pPATI(不含patatin部分信号顺序)。pPATI通过基因枪介导在块茎切片中获得了瞬间表达。以上建构物通过农杆菌介导转入了马铃薯品种Desiree。X-Gluc染色(PATIs不能染色)及PCR结果证实已获得转基因植株。利用离体块茎诱导系统,GUS的表达进一步用荧光进行定量检测,结果显示,PATI-GUS的转基因植株中GUS比活性均以块茎明显高于茎段,达10-20倍。蔗糖浓度的升高,PATI-GUS植株中的GUS比活性无明显变化,与前人报道有不同。此外,光照促进PATI-GUS的表达。     

通过DNA改组技术获得高活性β-葡萄糖苷酸酶

β 葡萄糖苷酸酶是在植物转基因中广泛应用的报告基因 .以质粒pBI12 1中的GUS基因为基础 ,利用DNA改组方法 ,经DNaseⅠ降解 ,PrimerlessPCR ,PrimerPCR对GUS基因进行了突变和改组 ,然后将改组的GUS基因连接到原核表达载体pG2 5 1中 ,构建了库容为 10 8的突变体库 .经过活性的筛选 ,得到活性提高的克隆 ,再以此为基础 ,经过新的改组、筛选得到活性大幅度提高的克隆GUS2 4 .基因测序显示 ,GUS2 4与GUS基因之间的同源性为 99 7% ,共有 6个核苷酸位点发生了改变 ,分别是 :379位的A突变为G ,396位的T突变为C ,711位的G突变为A ,95 8位T突变为C ,990位的T突变为C ,1649位的A突变为G .核苷酸序列推导的氨基酸序列显示 ,3个氨基酸发生了突变 ,12 7位的Ser突变为Gly ,32 0位的Trp突变为Arg ,5 5 0位的Asn突变为Ser.X gluc染色检测和荧光测活结果显示GUS2 4基因表达的 β 葡萄糖苷酸酶基较GUS基因表达产物活性提高 3倍     

Cloning and characterization of a cathepsin D inhibitor gene from Solanum tuberosum L.

A DNA clone encoding a cathepsin D inhibitor CathInh was isolated from a potato genomic library using a CathInh cDNA as hybridization probe. The amino acid sequence of the coding region is nearly identical with a CathInh cDNA and CathInh proteins previously isolated from a tuber-specific cDNA library and from tubers, respectively. Analysis of GUS activity resulting from expression of chimeric CathInh promoter-GUS genes in transgenic potato plants revealed expression exclusively confined to potato tubers. No GUS activity could be detected in any other organ of the transgenic plants either constitutively or after wounding or treatment with abscisic and jasmonic acid (JA). Interestingly, part of the promoter region of the CathInh gene, essential for GUS activity in tubers, shows striking similarity to promoter regions of tuber-specific class I patatin genes.     

Expression of a chimaeric granule-bound starch synthase-GUS gene in transgenic potato plants

Granule-bound starch synthase is the key enzyme in amylose synthesis. The regulation of this gene was investigated using a chimaeric gene consisting of a 0.8 kb 5 upstream sequence of the granule-bound starch synthase gene from potato and the -glucuronidase gene which was introduced into potato using an Agrobacterium tumefaciens binary vector system. The chimaeric gene was highly expressed in stolons and tubers, whereas the expression in leaves, stems or roots from greenhouse-grown plants was relatively low. However, leaves from in vitro grown plantlets exhibited an elevated GUS expression. The expression of the chimaeric gene was inducible in leaves by growth on relatively high concentrations of sucrose, fructose and glucose and was about 30- to 50-fold higher than in leaves from greenhouse-grown plants. The granule-bound starch synthase gene is expressed organ-specifically since stolons and tubers showed GUS activities 125- to 3350-fold higher than in leaves. The activities in these two organs are 3- to 25-fold higher than the expression of the CaMV-GUS gene. Histochemical analysis of different tissues showed that only certain regions of leaves and roots express high GUS activities. Stolons and tubers show high expression.     

A promoter directing high level expression in pistils of transgenic plants

The promoter of the potato (Solanum tuberosum L.) SK2 gene, encoding a pistil-specific basic endochitinase, was cloned. Various fragments of the SK2-promoter, from 1 kb down to 0.23 kb in length, were fused to the GUS reporter gene. Chimaeric SK2 promoter-GUS fusion constructs were transformed into potato by Agrobacterium tumefaciens-mediated transformation. The SK2-GUS transgenic potato plants exhibited a highly specific GUS activity in the pistil. Expression in the pistil was shown to be developmentally regulated. In addition to the GUS activity in pistils, transgenic plants also showed a much weaker ectopic expression in anthers. In other tissues no systematic expression was detectable. All SK2 promoter fragments analysed conferred pistil-specific expression without significant qualitative or quantitative differences, demonstrating that the regulatory elements mediating this expression pattern are located within a 230 bp SK2 promoter fragment. The SK2 promoter may be used to engineer high levels of expression in pistils of transgenic plants.