Identification of a Novel Polymorphism in Bovine lncRNA ADNCR Gene and Its Association with Growth Traits

Adipocyte differentiation-associated long noncoding RNA (ADNCR) is a newly discovered lncRNA. It plays function by targeting miR-204 to significantly regulates the expression of the target SIRT1 gene in preadipocytes both at the level of mRNA and protein, thereby inhibiting adipogenesis. The tetra-primer amplification refractory mutation system PCR (T-ARMS-PCR) strategy is fast and accuracy at a negligible cost for SNP genotyping in large samples. In the study, a novel SNP g.1263T>A in intron 1 of bovine ADNCR gene was found. Herein, the T-ARMS-PCR assay was applied to detect the genotypes of the novel SNP of bovine ADNCR gene in 1017 individuals from seven cattle breeds and validated the accuracy by DNA sequencing assay of ninety animals representing three different genotypes. The concordance between two different methods was 100%. The association analysis indicated that this locus was significantly associated with the body weight (P?=?0.010), chest girth (P?=?0.014) and rump length (P?=?0.038) in Jinnan cattle, hucklebone width (P?=?0.032) in Qinchuan cattle, the cannon circumference (P?=?0.019) in Jinjiang cattle, respectively. These novel findings may be used for marker-assisted selection (MAS) and contribute to the performance of beef cattle in the future.     

NMDA Receptor Heterogeneity During Postnatal Development of the Rat Brain: Differential Expression of the NR2A, NR2B, and NR2C Subunit Proteins

Abstract: Changes in the expression of the NMDA receptor subunits (NRs) NR2A, 2B, and 2C were investigated in histo blots of the developing rat brain with subunit-specific antisera. At birth, the NR2B subunit was detected almost ubiquitously, the NR2A subunit staining was faint and restricted to the hippocampus, cerebral cortex, and striatum, and no NR2C subunit immunoreactivity was detected. During the first 3 postnatal weeks, the NR2B subunit became confined to forebrain structures, whereas the NR2A immunoreactivity became abundantly expressed throughout the brain. The NR2C immunoreactivity emerged 5 days after birth in the olfactory bulb, thalamus, and vestibular nuclei and became very intense after 10 days in cerebellar granule cells, its primary site of expression in adulthood. After 3 weeks, NR2A and NR2B immunoreactivity decreased to adult levels, whereas NR2C immunoreactivity remained unchanged. The patterns of distribution of the subunit proteins were in agreement with those of their corresponding mRNAs, as monitored by in situ hybridization histochemistry, although the mRNA translation appeared to be delayed by several days in certain areas. Our results reveal a progressive increase in the heterogeneity of NMDA receptors due to the comparably late onset of NR2A and NR2C subunit expression and by the area-specific rearrangement of NR2B subunit expression following birth.     

PRPS2基因启动子区的单核苷酸多态性(英文)

人类基因组变异主要以单核苷酸多态性 (SNPs)为主 .采用多荧光标记的PCR单链构象多态性分析法 (MF PCR SSCP)对磷酰核糖焦磷酸合成酶亚基Ⅱ (PRPS2 )基因启动子区的序列进行了SNP的筛选 ,对筛选到的阳性片段进行序列分析以确定SNP的类型及位置 .在 1 5kb的启动子区发现了 2个SNP (- 10 79t c ,- 1110a g) .研究结果为PRPS2基因SNPs的数据库提供了新的信息 .     

生后早期听觉剥夺、经验改变大鼠听皮层NMDA受体NR2B蛋白表达

应用蛋白质印迹检测技术,研究早期听觉剥夺、经验对大鼠听皮层NMDA受体NR2B蛋白表达的影响.结果表明,听觉剥夺使生后14天龄组和28天龄组动物听皮层NR2B蛋白表达水平明显下降(P<0.05,P<0.01),听觉剥夺7天后再给予纯音暴露则又使NR2B表达水平明显提高(P<0.05),呈现双向调节趋势.听觉剥夺和纯音暴露对生后42天龄组大鼠听皮层NR2B表达不再产生明显调节作用(P>0.05).结果提示,在大鼠生后发育关键期,听觉剥夺、经验可改变听皮层NMDA受体NR2B蛋白表达水平.研究结果为研究感觉功能发育可塑性的机制提供了重要实验资料.     

E-钙粘蛋白基因启动子区域单核苷酸基因多态性(-347G→GA)和大肠癌遗传易感性的联系

目的:探讨E-钙粘蛋白基因(CDH1)启动子区域-347G→GA单核苷酸基因多态性(single nucleotide polymorphism,SNP)和中国地区散发大肠癌(colorectal cancer,CRC)人群遗传易感性的联系.方法:以中国地区大肠癌病人群为研究基础,同时建立无肿瘤家族史以及其它遗传性疾病史的正常对照组,进行病例对照研究,采用聚合酶链反应一限制性片段长度多态性技术(poly-merase chain reaction-restriction fragment length polymorphism,PCR-RFLP)行目的基因的多态性测定.结果:正常对照组和大肠癌组的目的基因型分布频率并无统计学差异(x2检验,P=0.246),而近端大肠癌患者中GA基因型(GA/GA杂合子和GA/GA纯合子)比例明显高于对照组(OR=1.773,95%CI=1.174-2.679,P=0.007),同时低分化大肠腺癌GA基因型比例也明显高于对照组(OR=2.623,95%CI=1.326-5.191,P=0.007).结论:CDHl启动子区域-347G→GA单核苷酸基因多态性中GA基因型可能增加近端大肠癌以及低分化腺癌的发病风险.     

人肝癌细胞LASP1 启动子的克隆及其核心调控区域的初步鉴定

目的:克隆人肝癌细胞的LASP1基因的启动子区域并找出该基因启动子的核心调控区域。方法:提取肝癌Hep G2细胞总DNA,PCR扩增不同长度的LASP1启动子片段,克隆至PGL3-Basic载体中构建重组表达载体PGL3-P1(2059 bp)、PGL3-P2(1123bp)、PGL3-P3(909 bp)、PGL3-P4(574 bp)和PGL3-P5(159 bp),转化入大肠埃希菌(E.coli)DH5α中,提取质粒经双酶切、PCR及测序鉴定阳性克隆并测序。将构建的重组表达载体、PGL3-Basic载体分别与内参质粒PRL-Tk共转染Hep G2细胞,48 h后经双荧光素酶报告基因检测试剂盒检测其活性。结果:PCR结果、双酶切结果以及DNA测序结果表明成功构建了LASP1启动子荧光素酶报告基因载体;双荧光素酶报告基因检测结果显示,与PGL3-Basic组相比,重组载体PGL3-P1、PGL3-P2、PGL3-P3、PGL3-P4均具有较强的启动子活性(P0.01),其中,PGL3-P4的活性最强。结论:成功构建了人肝癌细胞不同截断长度的LASP1基因启动子荧光素酶报告基因载体,确定了LASP-1基因启动子的核心区域(-581 bp~-8 bp),为进一步研究LASP1基因在肝癌细胞中表达的关键调节因素及分子机制奠定了基础。     

水稻液泡ATPase B 亚基基因的克隆及其在低磷胁迫下的表达特征分析

利用抑制性扣除杂交(SSH)技术构建水稻(Oryza sativa L.)根系磷饥饿诱导cDNA文库,获得编码液泡ATPase (V-ATPase) B亚基的克隆,通过反转录PCR方法获得该基因的完整序列.该基因编码487个氨基酸,含有一个保守的ATP结合位点,其蛋白分子量为54.06 kD,等电点为4.99.Southern印迹表明,V-ATPase B亚基基因在水稻基因组中以单拷贝形式存在.氨基酸同源性分析发现,V-ATPase B亚基是一个较为保守的蛋白亚基,其序列变化伴随生物的进化过程同步进行.Northern印迹表明,V-ATPase B亚基在水稻根系中受到磷饥饿诱导表达,磷饥饿6～12 h出现表达高峰,而在叶片中表达高峰有所滞后(24～48 h).在缺磷环境条件下,ATPase B亚基可能通过提高其表达量,进而提高质子转运活性,形成跨膜的电化学梯度,为体内储备磷跨液泡膜运输提供能量,从而提高植物体内磷的利用效率及其耐低磷的能力.     

水稻液泡ATPaseB亚基基因的克隆及其在低磷胁迫下的表达特征分析(英)

利用抑制性扣除杂交 (SSH)技术构建水稻 (OryzasativaL .)根系磷饥饿诱导cDNA文库 ,获得编码液泡ATPase (V_ATPase)B亚基的克隆 ,通过反转录PCR方法获得该基因的完整序列。该基因编码 4 87个氨基酸 ,含有一个保守的ATP结合位点 ,其蛋白分子量为 5 4 .0 6kD ,等电点为 4 .99。Southern印迹表明 ,V_ATPaseB亚基基因在水稻基因组中以单拷贝形式存在。氨基酸同源性分析发现 ,V_ATPaseB亚基是一个较为保守的蛋白亚基 ,其序列变化伴随生物的进化过程同步进行。Northern印迹表明 ,V_ATPaseB亚基在水稻根系中受到磷饥饿诱导表达 ,磷饥饿 6～ 12h出现表达高峰 ,而在叶片中表达高峰有所滞后 (2 4～ 4 8h)。在缺磷环境条件下 ,ATPaseB亚基可能通过提高其表达量 ,进而提高质子转运活性 ,形成跨膜的电化学梯度 ,为体内储备磷跨液泡膜运输提供能量 ,从而提高植物体内磷的利用效率及其耐低磷的能力。     

A Naturally Occurring Null Variant of the NMDA Type Glutamate Receptor NR3B Subunit Is a Risk Factor of Schizophrenia

Hypofunction of the N-methyl-D-aspartate type glutamate receptor (NMDAR) has been implicated in the pathogenesis of schizophrenia. Here, we investigated the significance of a common human genetic variation of the NMDAR NR3B subunit that inserts 4 bases within the coding region (insCGTT) in the pathogenesis of schizophrenia. The cDNA carrying this polymorphism generates a truncated protein, which is electrophysiologically non-functional in heterologous expression systems. Among 586 schizophrenia patients and 754 healthy controls, insCGTT was significantly overrepresented in patients compared to controls (odds ratio = 1.37, p = 0.035). Among 121 schizophrenia patients and 372 healthy controls, genetic analyses of normal individuals revealed that those carrying insCGTT have a predisposition to schizotypal personality traits (F_1,356 = 4.69, p = 0.031). Furthermore, pre-pulse inhibition, a neurobiological trait disturbed in patients with schizophrenia, was significantly impaired in patients carrying insCGTT compared with those with the major allele (F_1,116 = 5.72, p = 0.018, F_1,238 = 4.46, p = 0.036, respectively). These results indicate that a naturally occurring null variant in NR3B could be a risk factor of schizophrenia.     

大鼠Y染色体探针的制备与鉴定

目的:研究制备地高辛标记的大鼠性别决定基因Y区(Y染色体,SRY)探针,用于检测雄性大鼠来源的细胞在雌性受鼠体内的SRY基因表达情况.方法:按已知的雄性大鼠Y染色体上性别决定基因(SRY)的序列,请上海博亚公司合成oligoDNA,采用PCR技术连接并扩增,地高辛标记的方法制备基因探针.以雌性大鼠为对照,原位杂交法检测大鼠肾组织切片Y染色体阳性细胞情况.结果:用原位杂交法证实在雄性大鼠肾脏内有SRY表达,而雌性大鼠肾脏无Y染色体阳性细胞,证实这种探针具有较高的敏感性和特异性.结论:大鼠性别决定基因SRY探针的制备成功,为进一步研究异体雄性大鼠细胞移植后的分布和表达提供了实验基础.     

LASS1基因克隆及其在大鼠脑皮层的表达与神经元衰老的相关性初步研究

长寿保障基因LAG1是从酵母中克隆的与酵母寿命相关的基因,随酵母生命衰老而表达发生变化.对大鼠中同源基因LASS1进行克隆、测序和序列分析,发现其mRNA序列不同于GenBank中的预测序列,开放阅读框包含1 053碱基对,编码蛋白由350个氨基酸组成,内含Lag1蛋白家族保守的Lag1p motif和TLC结构域.从新生、1月龄、6月龄、12月龄和24月龄大鼠脑顶叶皮质提取总RNA,用半定量RT-PCR及RNA印迹方法对LASS1在大鼠脑皮质中的表达随年龄变化情况进行分析.结果表明,出生后LASS1表达量随年龄增加而增高,至6月龄达高峰,然后随年龄增加而逐渐下降,至24月老龄鼠达最低.衰老相关β半乳糖苷酶(SA-β-gal)对鼠脑皮层染色发现,神经元阳性染色随年龄增长明显增加.大鼠LASS1基因表达在正常衰老过程中发生变化,为进一步研究该基因的作用奠定了基础.     

Identification and Transgenic Analysis of a Murine Promoter that Targets Cholinergic Neuron Expression

Abstract : Choline acetyltransferase (ChAT) is a specific phenotypic marker of cholinergic neurons. Previous reports showed that different upstream regions of the ChAT gene are necessary for cell type-specific expression of reporter genes in cholinergic cell lines. The identity of the mouse ChAT promoter region controlling the establishment, maintenance, and plasticity of the cholinergic phenotype in vivo is not known. We characterized a promoter region of the mouse ChAT gene in transgenic mice, using β-galactosidase ( LacZ ) as a reporter gene. A 3,402-bp segment from the 5'-untranslated region of the mouse ChAT gene (from -3,356 to +46, +1 being the translation initiation site) was sufficient to direct the expression of LacZ to selected neurons of the nervous system ; however, it did not provide complete cholinergic specificity. A larger fragment (6,417 bp, from -6,371 to +46) of this region contains the requisite regulatory elements that restrict expression of the LacZ reporter gene only in cholinergic neurons of transgenic mice. This 6.4-kb DNA fragment encompasses 633 bp of the 5'-flanking region of the mouse vesicular acetylcholine transporter (VAChT), the entire open reading frame of the VAChT gene, contained within the first intron of the ChAT gene, and sequences upstream of the start coding sequences of the ChAT gene. This promoter will allow targeting of specific gene products to cholinergic neurons to evaluate the mechanisms of diseases characterized by dysfunction of cholinergic neurons and will be valuable in design strategies to correct those disorders.     

多巴胺D4受体基因启动子区多态性与精神分裂症的相关性

目的:探讨多巴胺D4受体(Dopaminc D4 receptor,DRD4)基因启动子区的3个功能多态性与精神分裂症是否存在相关性.方法:严格按照诊断标准,选取无亲缘关系的精神分裂症患者220例,健康对照组200例提取基因组DNA,采用聚合酶链反应及等位基因特异性扩增技术检测DRD4基因启动子区-521C/T、-616C/G和-1240L/S 3个功能位点的基因型,采用HaploView4.0及SPSS11.5软件分析各位点基因型、等位基因频率及组间差异.结果:DRD4基因-1240L/S的基因型及等位基因频率分布在精神分裂症与正常对照组存在显著性差异(p<0.05).DRIM基因启动子区-521C/T和-616C/G位点的基因型及等位基因频率分布在精神分裂症组与正常对照组无统计学差异(p>0.05).结论:DRD4基因-1240L/S多态性与精神分裂症相关联,携带有-1240L/S多态性住点L等住基因的个体可能更容易患精神分裂症.