The cloning and preliminarily functional analysis of the human neurotrimin gene

~~The cloning and preliminarily functional analysis of the human neurotrimin gene~~     

DNA片段的高效克隆

用Taq酶进行PCR扩增时,其PCR产物的3末端有一个附加的A碱基。因此,目前在克隆PCR产物时一般使用T－VCctor。但T—Vector的价格比较昂贵,而使用本试剂盒也可在短时间内使PCR产物与平滑末端载体进行高效连接。PCR产物与平滑末端的载体连接时,有必要除去3廉端的附加碱基,并使其5末端磷酸化。使用TaKaRaBKLKit（BllllltillgKillstiollLigstiollKit）可使这一连串的反应在短时间内完成。PCR产物的末端平滑化与磷酸化反应在一个反应体系内同时进行,一次反应后便可得到能够用于连接的DNA片段。用于连接反应的PCR产物无需进…     

多重实时荧光PCR相对定量法快速诊断唐氏综合征

为了建立一种基于多重实时荧光相对定量PCR技术并应用之于唐氏综合征分子诊断, 选择21号染色体上唐氏综合征特异区域基因片段(DSCR3)为目的基因, 以12号染色体上的磷酸甘油醛脱氢酶基因(GAPDH)为参照基因, 设计合成两对引物以及分别以不同荧光标记的TaqMan探针, 在同一个反应管中进行扩增。以相对定量指标△CT值区分唐氏综合征患者与正常人。采用EB 病毒转化技术, 把唐氏综合征患者外周血B 淋巴细胞转化成永生淋巴母细胞系作为标准品。通过优化反应条件, 使得目的基因和参照基因的扩增效率基本一致, 接近100%, 模板浓度在3～300 ng/μL范围内, △CT值的变异系数小于15%, 浓度在30 ng/μL时, 变异系数最小(＜10%), 以该浓度的DNA作为模板进行批内和批间实验的△CT值重复性好, 变异系数分别为9.8%和13.3%。运用建立的方法检测20例唐氏综合征患者的血标本和30例正常人的血标本, 正常人△CT值范围是-1.90～-1.30, 患者的△CT值范围是-2.95～-2.15, 两组之间无交叉重叠, 有明显差异(P＜0.001)。唐氏综合征患者永生细胞系建系成功 ,染色体核型和DNA 分析表明建系前后遗传是稳定的。因此, 实时荧光定量PCR比较△CT值的相对定量法快速诊断唐氏综合征是可行的。     

Phylogenetic Relationships of the Genus Paramesotriton （Caudata： Salamandridae） with the Description of a New Species from Qixiling Nature Reserve,Jiangxi, Southeastern China and a Key to the species

The matrilineal genealogy of the genus Paramesotriton is hypothesized based on DNA sequences from mitochondrial NADH subunit two （ND2） and its flanking tRNAs （tRN^Arp and a partial tRNA^Ala）. The genealogy identifies a highly divergent, unnamed lineage from Qixiling Nature Reserve, Jiangxi, China and places it as the sister taxon of P. chinensis. The newly discovered population differs from other congeners by several features of external morphology including having large clusters of dark brown conical warts on the dorsum of the head, lateral surface of the body and dorsolateral ridges. Its intermittent dorsal vertebral ridge is the same color as other parts of the dorsum and tail narrows gradually from the base to the tip. Further, the new population differs from all congeners by an uncorrected P-distance of more than 9.38% in ND2. Consequently, we describe the new species of Asian warty newt （Salamandridae） as Paramesotriton qixilingensis sp. nov.     

Effects of MEK inhibitor U0126 on meiotic progression in mouse oocytes： microtuble organization, asymmetric division and metaphase Ⅱ arrest

In this study we used U0126, a potent and specific inhibitor of MEK, to study the roles of MEK/ERK/p90~(rsk) signaling pathway in the meiotic cell cycle of mouse oocytes. The phosphorylation of MAP kinase and p90~(rsk) in the oocytes treated with 1.5 μM U0126 was the same as that in oocytes cultured in drug-free medium. With 1.5 μM U0126 treatment, the spindles appeared normal as they formed in oocytes, but failed to maintain its structure. Instead, the spindle lost one pole or elongated extraordinarily. After further culture, some oocytes extruded gigantic polar bodies (>30 μm) that later divided into two small ones. Some oocytes underwent symmetric division and produced two equal-size daughter cells in which normal spindles formed. In oocytes with different division patterns, MAP kinase was normally phosphorylated. When the concentration of U0126 was increased to 15 mM, the phosphorylation of both MAPK and p90~(rsk) were inhibited, while symmetric division was decreased. When incubating in medium c     

Identification of γ_1 subunit of GABA_A receptor in rat testis

INTRODUCTIONry-Aminobutyric acid (GABA) is the predomi-nant inhibitory neurotransmitter in the vertebratecentral nervous system (CNS)[1]. Whereas outsidethe CNS, many peripheral tissues have also beenfOund to have GABAergic system[2].The mammalian sperm acrosome reaction (AR)is a modified exocytotic event that is essential to thefertilization process[3]. Two main agOnists of AR,the zona pellucida glycoprotein ZP3[4] and proges-terone[5], have been identified in the oocyte vest-me…     

γ-氨基丁酸对鳜摄食和食欲的影响

为阐明γ-氨基丁酸(GABA)对鳜(Siniperca chuatsi)摄食和食欲的影响, 对鳜脑室注射生理盐水和不同剂量的GABA(50、125、500和2000 μg)。结果显示, 注射125 μg GABA组的鳜在2h内摄食量显著升高。实时荧光定量PCR (RT-qPCR)结果显示, 注射125 μg GABA 0.5h后, 鳜鱼脑中AgRP和NPY mRNA表达量上调, CART和POMC mRNA表达量下调, 都和鳜摄食量增加相一致。相比于对照组, 注射GABA后Leptin-R的mRNA表达量在0.5h和2h都有显著下降。这些结果表明GABA可能通过leptin的信号通路来影响食欲, 进而影响摄食量。研究结果可以为GABA在水产饲料中的应用提供理论依据。     

Phylogenetic relationship of 16 Oedipodidae species （Insecta： Orthoptera） based on the 16S rRNA gene sequences

The sequences of the mitochondrial 16S rRNA gene of 16 Oedipodidae species were amplified and sequenced. All sequences were aligned and analyzed and the phylogenetic relationships were inferred. The properties of 16S gene in Oedipodidae showed typical patterns of many insects such as a high A＋T content and variable distance-dependent transition/transversion ratios. The 0.2 weight for sites of loops may be advisable for phylogeny reconstruction using the maximum parsimony method. The phylogenetic analysis results do not support the current subfamily classification systems of Oedipodidae. Bryodemellinae and Bryodeminae are closely related and should be merged as one subfamily. The status of Oedipodinae and Locustinae is also problematic.     

Temporal dynamics of mental impasses underlying insight-like problem solving

Insight problem solving is characterized by mental impasses,states of mind in which the problem solver does not know what to do next.Although many studies have investigated the neural correlates of insight problem solving,however,the question when mental impasses occur during insight problem solving has been rarely studied.The present study adopted high temporal resolution ERPs to investigate the temporal dynamics of an impasse underlying insight problem solving.Time locked ERPs were recorded associated with problems with impasses(PWI) and problems without impasses(POI).The problem types were determined by participants’ subjective responses.The results revealed an early frontocentral P2 was linked with the preconscious awareness of mental impasses and a P3a was associated with fixed attention when the impasse formed.These findings suggest the impasse may occur initially at a relatively early stage and metacognition plays an important role in insight problem solving.     

The genetic equidistance result: misreading by the molecular clock and neutral theory and reinterpretation nearly half of a century later

In 1963,Margoliash discovered the unexpected genetic equidistance result after comparing cytochrome c sequences from different species.This finding,together with the hemoglobin analyses of Zuckerkandl and Pauling in 1962,directly inspired the ad hoc molecular clock hypothesis.Unfortunately,however,many biologists have since mistakenly viewed the molecular clock as a genuine reality,which in turn inspired Kimura,King,and Jukes to propose the neutral theory of molecular evolution.Many years of studies have found numerous contradictions to the theory,and few today believe in a universal constant clock.What is being neglected,however,is that the failure of the molecular clock hypothesis has left the original equidistance result an unsolved mystery.In recent years,we fortuitously rediscovered the equidistance result,which remains unknown to nearly all researchers.Incorporating the proven virtues of existing evolutionary theories and introducing the novel concept of maximum genetic diversity,we proposed a more complete hypothesis of evolutionary genetics and reinterpreted the equidistance result and other major evolutionary phenomena.The hypothesis may rewrite molecular phylogeny and population genetics and solve major biomedical problems that challenge the existing framework of evolutionary biology.