脂蛋白脂酶基因的研究进展

脂蛋白脂酶(lipoprotein lipase, LPL)是脂质代谢的关键酶, 主要催化乳糜微粒和极低密度脂蛋白中的甘油三酯水解, 产生供组织利用的脂肪酸和单酰甘油。LPL基因突变影响LPL活性, 导致脂质代谢紊乱, 与2型糖尿病、高血压、动脉硬化、肥胖、冠心病的发病风险相关联。文章综述了LPL基因的结构、功能、表达调控以及与复杂疾病的关联研究进展。     

脂蛋白脂酶基因与apo C—Ⅱ基因变异

原发性高甘油三酯血症是由于血浆中的甘油三酯持续升高,达到了一个高水平而赞成的一组症侯,甘油三酯代谢紊乱的病人往往带有家族性,提示某些基因的遗传缺陷可能是导致高甘油三酯血症的原因之一,具有常染色体隐性遗传特性,引起家族性乳糜微粒血症的两个主要因素是脂蛋白脂酶（LPL）以及LPL的协同因子载脂蛋白C－Ⅱ（apo C－Ⅱ）的异常,对于这些基因变异目前已经有了比较成熟的检验方法,但是基因治疗的治疗尚处在研究阶段。     

脂蛋白脂酶基因与代谢综合征关系的研究进展

脂蛋白脂酶是脂质代谢的关键酶之一,是血浆中清除甘油三酯的限速酶,主要催化乳糜微粒和极低密度脂蛋白核心中的甘油三酯水解,使得机体能够利用由食物摄取和肝脏合成的脂肪,同时释放出脂肪酸和单酰甘油.脂蛋白脂酶基因突变可以影响脂蛋白脂酶的活性,从而导致脂代谢紊乱,与肥胖、胰岛素抵抗、2型糖尿痛和高血压的发病风险相关.本文综述了脂蛋白脂酶的结构、功能、基因结构及其多态性与血脂和代谢综合征关系的研究进展.     

脂蛋白脂酶基因多态性与2 型糖尿病的相关性研究

目的：探讨脂蛋白脂酶（lipoprotein lipase,LPL）基因PvuⅡ酶切多态性与2型糖尿病的相关性。方法：采用聚合酶链反应-限制性片段长度多态性（PGR-RFLP）方法,分析了156例样本LPL基因第6内含子PvuⅡ多态性（病例组98人。对照组58。其中40个2型糖尿病同胞对,病例组40人,对照组40人）。结果：病例组与对照组的基因型和基因频率均无显著性差异。结论：湖北汉族人群脂蛋白脂酶基因PvuⅡ酶切多态性与2型糖尿病无明显关联。     

脂蛋白脂酶基因HindⅢ酶切多态性与2型糖尿病的关联研究

为了探讨脂蛋白脂酶(lipoprotein lipase, LPL)基因HindⅢ酶切多态性与2型糖尿病(type 2 diabetes mellitus, T2DM)的关系, 采用病例-同胞对照设计和随机病例-对照设计, 应用PCR-RFLP方法, 对264例T2DM患者和102名正常人LPL基因HindⅢ酶切多态性进行分析。结果表明, T2DM组H+等位基因及H+H+基因型的频率较对照组显著增高(H+: 76.9%比69.1%, P<0.05; H+H+: 59.8%比52%, P<0.05)。根据实验设计分组, 同胞对T2DM组H+等位基因及H+H+基因型的频率较同胞对对照组显著增高(H+: 81.5%比67.8%, P<0.05; H+H+: 68.5%比50.7%, P<0.05), 而随机病例组与对照组间无此频率差异性(P>0.05)。多因素Logistic回归显示T2DM的独立危险因素是空腹血糖和LPL基因型, H+H+ 纯合子患T2DM的危险性是H+H-和H-H-基因型的1.995倍(95% CI: 1.036～3.840, P<0.05)。提示LPL基因HindⅢ多态性与湖北汉族人T2DM的危险性相关, 其中H+等位基因可能是T2DM的遗传危险因素。     

鳜脂蛋白脂酶基因SNP及其与食性驯化相关性分析

鳜食性奇特,通常情况下拒食死饵或配合饲料,在长期的养殖过程中发现:通过驯养,能逐步诱导部分鳜以非活饵为食。通过分子标记定向选育易驯化鳜并使用人工饲料大规模养殖,可以有效解决鳜养殖中存在的成本高、污染严重、病害严重等问题。脂蛋白脂酶基因(Lipoprotein lipase LPL)是脂蛋白代谢的关键酶之一,生理功能是将乳糜微粒和极低密度脂蛋白核心的甘油三酯催化分解为甘油和脂肪酸,以供组织氧化供能和贮存。文章采用PCR产物直接测序法对鳜脂蛋白脂酶基因6、7内含子和6、7、8外显子进行了SNPs遗传多态性检测和分析,探寻LPL基因的等位基因及其基因型在两个食性驯化表型群体中的分布情况。结果在第7外显子处共检测到3个SNPs位点(A25T、G26T和C29G),其中A25T、C29G两个为非同义突变。利用卡方检验分析驯化与未驯化组,结果表明LPL基因3个SNPs位点对鳜的食性驯化不具显著差异性(P>0.05)。将3个SNPs位点不同基因型组合成5种双倍型,卡方检验表明双倍型Dip2在两组中存在显著差异(P<0.05)。文章成功完成鳜LPL基因组部分区段多态性分析,因而可以考虑将LPL基因作为影响鳜食性驯化的候选基因,作为遗传标记,为今后的标记辅助选择育种工作奠定基础,具有广泛的应用前景。     

脂蛋白脂酶与肥胖

脂蛋白脂酶(lipoprotein lipase,LPL)是甘油三酯分解代谢的限速酶,与肥胖发生联系密切.综述了LPL蛋白的分子结构特点、分泌方式,及其参与的与肥胖发生有关的脂肪细胞分化、脂质沉积、脂代谢等过程.     

11例重度内源性高甘油三酯血症患者脂蛋白脂酶基因…

对成都地区１１例重度内源性高甘油三酯血症患者脂蛋白脂酶基因外显子１－９的序列进行了分析。结果发现，１１例患者中有４例其ＬＰＬ基因在外显子６，８及９具有３种杂合子变异。其中一种为ｈｒ＾３６１－Ｔｈｒ（外显子８，Ｃ＾１３３８→Ａ）突变，为无义突变，尚未见报道，在７４例血脂正常中国人中的发生频率为１１．４％。另外２种变异Ａｌａ＾３６１－Ｔｈｒ（外显子６，Ｇ＾１０３７→Ａ）及Ｓｅｒ＾４４７－Ｔｅｒ外显子９     

脂蛋白脂酶基因的克隆、序列测定及定点突变

 以人的脂肪组织总RNA为模板 ,参考已报道的脂蛋白脂酶 (lipoproteinlipase ,LPL)cDNA设计引物 ,利用RT PCR方法扩增得到了LPLcDNA ,并经序列测定证实其序列是正确的 .在冠心病患者LPL基因第 5外显子的 830位碱基处发现了G→A的转换 ,该变异导致LPL基因第 192位的密码子CGA被CAA取代 ,使LPL第 192位精氨酸改变为谷氨酰胺 .在变异碱基附近设计合成两条引物 ,其中一条包含所要改变的碱基 ,利用基于PCR的定点突变技术和体外重组的方法获得了G830A变异的LPLcDNA     

苯丙氨酸羟化酶（PAH）基因外显子7及其两侧内含子的突变研究

为探讨中国苯丙酮尿症（PKU）人群中苯丙氨酸羟化酶（PAH）基因外显子7的突变特征,对147例PKU患儿的294个PAH基因外显子7以及两侧部分内含子序列,应用PCR－单链构象多态性（SSCP）分析及基因序列分析的方法进行了筛查和确定。共发现13种突变基因：G239D、R241C、R241fs、R243Q、G247S、G247V、R252Q、L255S、R261Q、M276K、E280G、P281L、Ivs7+2T>A,其中7 种突变基因在中国PKU人群首次发现：G239D 、R241fs 、G247S 、E280G、L255S、R261Q、P281L,前4种在国际上尚未见到报道,并已提交到国际PAH突变数据库（www.pahdb.mcgill.ca）。突变基因的总频率为30.61%（90 /294）。突变涉及了错义、缺失、移码和剪接位点4种突变类型。结果明确了PAH基因外显子7的突变种类和分布等特征,表明外显子7是中国人PAH基因突变的热点区域。 Abstract: To study mutation in exon 7 of the gene for the phenylalanine hydroxylase（PAH）, the mutations in exon 7 and flanking sequence of PAH gene were detected by means of SSCP analysis and DNA sequencing, in 147 unrelated Chinese children with phynelketonuria and their parents. Thirteen different mutations, including 11 missense, 1 deletion and 1 splice mutation, were revealed in 90/294 mutant alleles (30.61%). The prevalent mutations were R243Q (22.8%) and Ivs7nt2t->a (2.38%). Seven novel mutations were identified: G239D, R241fsdelG, G247S, E280G, L255S, R261Q, P281L. These new mutations have not been described in Chinese PKU population and the first 4 mutants have not been reported and thus been submitted to www.pahdb,mcgill.ca. The missense was the most common type. The deletion and frameshift mutations were detected for the first time in Chinese PKU population. This study showed the mutation characteristics and their distribution in exon 7 of PAH gene and proved that the exon 7 was the hot region of PAH gene mutation in Chinese PKU population .     

中国广东汉族人群核苷酸修复基因hMTH1 遗传多态性研究

为研究中国南方汉族人群核苷酸修复基因hMTH1遗传多态性,应用聚合酶链反应-单链构象多态性技术检测172名健康人外周血白细胞hMTH1基因启动子及全部5个外显子多态性,并进行DNA测序。结果发现hMTH1基因启动子及外显子1序列保守,未见突变;外显子2第73位碱基存在T→C杂合型突变,基因型TT和TC频率分别为93.02%、6.98%,等位基因T和C频率分别为96.51%、3.49％;外显子3第45位遗传密码存在T→C杂合型突变,基因型TT和TC频率分别为95.35%、4.65%,等位基因T和C频率分别为97.67%、2.33%,该多态性为首次发现;外显子4第83位遗传密码存在G→A杂合型突变,基因型GG和GA频率分别为89.53%、10.47%,等位基因G和A频率分别为94.77%、5.23％;外显子5第119位氨基酸遗传密码存在C→T杂合型突变,基因型CC和CT频率分别为95.93%、4.07%,等位基因C和T频率分别为97.97%、2.03％。Abstract: In order to study the genetic polymorphisms of nucleotide repair gene hMTH1 in southern Chinese Han population, the polymorphisms of the gene’s promoter and its five exons among peripheral blood lymphocytes of 172 Chinese Han people were analyzed with polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and DNA sequencing. The sequences of the promoter and exon 1 of hMTH1 gene were conserved. A T to C polymorphism was detected at the 73th base in exon2. The genotype frequencies of TT and TC were 93.02% and 6.98%, respectively. The allelic frequencies of T and C were 96.51% and 3.49％, respectively. A T to C polymorphism was detected at codon 45 in exon3, which was first reported. The genotype frequencies of TT and TC were 95.35% and 4.65%, respectively. The allelic frequencies of T and C were 97.67% and 2.33%, respectively. A G to A polymorphism was detected at codon 83 in exon4. The genotype frequencies of GG and GA were 89.53% and 10.47%, respectively. The allelic frequencies of G and A were 94.77% and 5.23％, respectively. A C to T polymorphism was detected at codon 119 in exon5. The genotype frequencies of CC and CT were 95.93% and 4.07%, respectively. The allelic frequencies of C and T were 97.97% and 2.03％, respectively.     

Small GTP-Binding Protein Gene Is Associated with QTL for Submergence Tolerance in Rice

Small GTP-binding proteins play critical roles in signal transduction in mammalian and plant systems. In this study, sequence variation of a small GTP-binding protein identified in the subgenomic region was analyzed. The major quantitative trait locus (QTL) controlling submergence tolerance on the 6.5-cM region of chromosome 9 was previously mapped, sequenced, and annotated. One of the most interesting candidate genes located in this QTL was a 5.2-kb sequence, which included a coding sequence consisting of two exons and a promoter. The deduced amino acid sequence corresponded to a 24.8 kD protein consisting of 226 amino acids, with 98% identity to RGP1, a small GTP-binding protein involved in a signal pathway responding to hormones, such as cytokinin and ethylene. According to the amino acid sequence, a putative small G-protein was classified as a small Ras-related GTP-binding protein. DNA gel blot analysis showed that the putative gene encoding the Ras-related GTP-binding protein was present as a single copy in the rice genome. Comparison of genomic sequences from several rice cultivars tolerant to submergence identified single nucleotide polymorphisms located in the TATA box of the Ras promoter region. Linkage analysis showed that the putative gene for GTP-binding protein was tightly linked to the peak of the QTL previously mapped on the long arm of chromosome 9. The single strand conformation polymorphism of the putative GTP-binding protein gene can be used for allele discrimination and marker assisted selection for tolerance to flash flooding.     

Heterogeneity of HLA-G Genes Identified by Polymerase Chain Reaction/Single Strand Conformational Polymorphism (PCR/SSCP)

A genomic HLA-G clone named 7.0E was isolated from a Japanese placenta. The deduced amino acid sequence of the 7.0E was identical to two HLA-G genomic clones and two cDNA clones previously described. The DNA sequences of α1 and α2 domains of the HLA-G gene from 5 cell lines also encoded the same amino acids. However, a 14 bp insertion, ATTTGTTCATGCCT, was present in the 3′ untranslated region of 7.0E compared with the originally described HLA-G clone (HLA 6.0). Polymerase chain reaction (PCR)/single strand conformational polymorphism (SSCP) analysis of exon 8 allowed the HLA-G gene to be classified into two alternative types, G6.0 and 7.0 E, those correlated to the absence or the presence of the 14 bp stretch. Each group had minor sequence variant(s), and the alleles of the 7.0E-type were more heterogeneous than those of the G6.0- type. The 14 bp deletion is present only in the G6.0-type of HLA-G alleles among HLA class I genes. Thus it was suggested that G6.0 alleles were generated after diversification of the HLA-G.     

DNA单链二级结构预测与单链构象多态性分析的一致性研究

单链构象多态性(SSCP)分析是一种简便,快速检测DNA突变的方法,它在基因突变检测、遗传分析、进化研究等领域有着广泛的应用价值.但是这种方法的突变检出率随DNA序列不同而变化,一般只能达到70%～80%.这主要是有的碱基突变对单链DNA的构象影响较小,不能通过SSCP检测出来.将计算机对DNA二级结构的预测结果和实验结果作了对比,发现二者有很高的一致性.这一结果表明计算机的DNA单链二级结构预测分析可用于PCR-SSCP分析的辅助设计,提高SSCP的突变检出率.     

毛细管电泳-单链构象多态分析检测胃癌中p16基因突变

目的:建立快速高效检测胃癌患者胃癌组织及癌旁正常组织中p16基因突变的方法。方法:采用PCR扩增p16基因第二外显子易发生突变片段,扩增样品纯化后经95℃变性;以毛细管电泳(CE)分析法结合单链构象多态性(SSCP)对60例胃癌患者p16基因突变情况进行分析。结果:分析结果表明只有3例低分化腺癌患者存在基因突变,测序表明p16基因第二外显子碱基序列AGAC发生碱基A丢失。结论:p16基因突变可能导致胃癌的发生,但不起主导作用;CE-SSCP分析方法具有快速、灵敏、准确的特点,可用于胃癌组织中p16基因的突变分析。     

豚鹿MHC-DOA基因克隆及其外显子2遗传多样性研究

主要组织相容性复合物(MHC)在脊椎动物的免疫系统中起着重要的作用,MHC-DOA基因是MHCⅡ类的非经典基因,其外显子2多态性丰富。豚鹿Axis porcinus是我国极度濒危的动物。本研究以成都动物园圈养的38头豚鹿为研究对象,提取豚鹿血液总RNA,反转录合成cDNA为模板,利用PCR方法克隆到豚鹿MHC-DOA序列;同时利用PCR方法扩增38头豚鹿的MHC-DOA基因外显子2基因并测序,再进行多态性分析。结果显示:豚鹿MHC-DOA基因开放阅读框长753 bp,编码250个氨基酸残基;蛋白质同源性分析表明,豚鹿MHC-DOA基因与东欧马鹿Cervus elaphus hippelaphus的同源性最高(98.4%);38头豚鹿的MHC-DOA外显子2共有10种单倍型,整体遗传多样性水平中等;MHC-DOA外显子2部分序列中发生了核苷酸的缺失和插入,进而引起DOA蛋白序列发生改变。豚鹿MHC-DOA多态性的研究对豚鹿种群遗传结构调查、遗传资源的保护具有重要意义。