利用修饰的随机引物构建非洲爪蟾酵母双杂交定向cDNA文库

描述了一种新的构建cDNA文库的方法,其中用来合成cDNA第一链的随机引物5'-端被加上碱基d(AC),在cDNA双链合成后所添加的连接头的末端含有碱基d(GTCG).在cDNA舣链3'-端,连接头连接上后会形成一个完整的Sall酶切位点d(GTCGAC),而在5'-端几乎不会形成Sall位点.在Sall酶切后利用形成的3'-粘性末端与5'-EcoRI粘性末端一起将cDNA双链定向导入线性化的质粒载体,再通过转化细菌获得cDNA文库.利用此方法构建了一个不同发育时期非洲爪蟾胚胎酵母双杂交cDNA文库.检测了文库中空载体的比例,插入片段大小和不同基因的表达水平几个指标,都符合预期,但是同时发现定位于mRNA的3'-端的插入片段比例比较低.这种倾向性符合文献报道,应该是实验系统的倾向性,不影响进一步的酵母双杂交实验.这些数据证明成功地构建了定向非洲爪蟾酵母双杂交cDNA文库.

Temporal and Spatial Expression Patterns of Sox1 Gene in Xenopus laevis

克隆了非洲爪蟾的Sox1基因并研究了它在非洲爪蟾早期发育过程中的时空表达图式,比较了Sox1-3基因在发育的脑和眼中的表达图式.序列比对分析显示Sox1-3蛋白在其HMG框结构域具有高度的保守性.通过RT-PCR方法分析了Sox1基因在爪蟾早期不同发育时段的表达情况,结果显示Sox1基因从未受精卵到尾芽期均有表达,但表达强度有所差异.原位杂交结果显示,在早期卵裂阶段和囊胚期,Sox1基因主要在动物极表达;从神经板期开始,Sox1基因主要在中枢神经系统和眼原基中表达.在蝌蚪期,Sox1与Sox2、Sox3在脑部和眼睛的表达区域有所不同.对于爪蟾Sox1基因时空表达图式的研究将有助于阐明SoxB1基因家族在脊椎动物神经系统发生过程中的作用.

文昌鱼RACK1基因在胚胎发育中的表达

脊椎动物活化的(蛋白)激酶C受体1(receptor for activated C-kinase 1,RACK1)在胚胎发育过程中起着至关重要的作用.克隆了RACK1基因在青岛文昌鱼(Branchiostoma belcheri)中的同源基因AmphiRACK1,对其进行了系统的进化学分析,并研究了该基因在正常胚胎发育过程中和经LiCl处理胚胎发育中的时空表达图式.系统进化学分析结果表明,文昌鱼RACK1位于脊椎动物进化枝的基部.在正常胚胎发育中,AmphiRACK1基因在脑泡、神经管和体节中都有明显的表达.在经LiCl处理的胚胎中,该基因在体节中的分节型表达变模糊,表达下调;在脑泡和神经管中的表达下调甚至消失.此外,在成体文昌鱼的轮器、鳃血管、肝盲囊和肠上皮以及精巢中都检测到AmphiRACK1基因不同程度的表达.

天然可降解脱钙骨基质材料复合脂肪干细胞的三维培养

背景:脱钙骨基质是一种常用的组织工程支架材料,脂肪干细胞是近年来发现的一种成体干细胞,二者均可自体取材,二者复合培养的研究还未见报道。目的:观察脂肪干细胞与脱钙骨基质复合后三维培养的生长、增殖情况。方法:无菌切取兔腹股沟皮下脂肪垫,采用Ⅰ型胶原酶搅拌消化法分离培养原代脂肪干细胞,系列传代传至第3代,倒置显微镜下观察细胞形态及生长情况,免疫荧光细胞化学染色检测细胞表面抗原的表达。切取兔髂骨,制备脱钙骨基质。取第3代脂肪干细胞,与脱钙骨基质复合后共培养,1周后扫描电镜观察细胞黏附和生长情况,2周后石蜡包埋切片进行苏木精-伊红染色。结果与结论:自兔皮下脂肪分离培养获得的脂肪干细胞增殖迅速,第3代脂肪干细胞表面抗原CD44呈阳性表达,CD34为阴性;制备的脱钙骨基质为三维立体多孔结构,具有良好的可塑性,并有一定的机械强度。复合培养后电镜扫描、苏木精-伊红染色显示,细胞在脱钙骨基质表面和孔隙内黏附生长良好,并能继续增殖和分泌胞外基质。结果提示脂肪干细胞与脱钙骨基质复合共培养生长增殖良好,并能分泌细胞外基质。

Cloning and developmental expression of the Xenopus Nkx6 genes

The evolutionarily conserved Nkx6 family transcription factors play important roles in the patterning of the central nervous system (CNS) and pancreas in vertebrates. In this study, we describe the cloning and expression patterns of the three Nkx6 family

Developmental expression of amphioxus RACK1

Vertebrate RACK1 plays a key role in embryonic development. This paper described the cloning, phylogenetic analysis and developmental expression of AmphiRACK1, the RACK1 homologous gene in amphioxus. Phylogenetic analysis indicated that amphioxus RACK1 wa

Embryonic stages and eye-specific gene expression of the local cyprinoid fish Anabarilius grahami in Fuxian Lake, China

Anabarilius grahami is a cyprinoid fish endemic to Fuxian Lake, Yunnan, China. In this study, a comprehensive staging series of A. grahami was produced. The embryonic development of A. grahami was divided into six main periods: zygote period, cleavage per

Islet-1 is required for ventral neuron survival in Xenopus

Islet-1 is a LIM domain transcription factor involved in several processes of embryonic development. Xenopus Islet-1 (Xisl-1) has been shown to be crucial for proper heart development. Here we show that Xisl-1 and Xisl-2 are differentially expressed in th

Xenopus RCOR2 (REST corepressor 2) interacts with ZMYND8, which is involved in neural differentiation

Regulation of neuronal gene expression is critical to nervous system development. REST (RE1-silencing transcription factor) regulates neuronal gene expression through interacting with a group of corepressor proteins including REST corepressors (RCOR). Here we show that Xenopus RCOR2 is predominantly expressed in the developing nervous system. Through a yeast two-hybrid screen, we isolated Xenopus ZMYND8 (Zinc finger and MYND domain containing 8) as an XRCOR2 interacting factor. XRCOR2 and XZMYND8 bind each other in co-immunoprecipitation assays and both of them can function as transcriptional repressors. XZMYND8 is co-expressed with XRCOR2 in the nervous system and overexpression of XZMYND8 inhibits neural differentiation in Xenopus embryos. These data reveal a RCOR2/ZMYND8 complex which might be involved in the regulation of neural differentiation. (C) 2010 Elsevier Inc. All rights reserved.

RNF220, an E3 ubiquitin ligase that targets Sin3B for ubiquitination

Modification of proteins by ubiquitination plays important roles in various cellular processes. During this process, the target specificity is determined by ubiquitin ligases. Here we identify RNF220 (RING finger protein 220) as a novel ubiquitin ligase for Sin3B. As a conserved RING protein, RNF220 can bind E2 and mediate auto-ubiquitination of itself. Through a yeast two-hybrid screen, we isolated Sin3B as one of its targets, which is a scaffold protein of the Sin3/HDAC (histone deacetylase) corepressor complex. RNF220 specifically interacts with Sin3B both in vitro and in vivo. Sin3B can be regulated by the ubiquitin-proteasome system. Co-expression of RNF220 promotes the ubiquitination and proteasomal degradation of Sin3B. Taken together, these results reveal a new mechanism for regulating the Sin3/HDAC complex. (C) 2010 Elsevier Inc. All rights reserved.

Differential expression of the Brunol/CELF family genes during Xenopus laevis early development

The BRUNOL/CELF family of RNA-binding proteins plays important roles in post-transcriptional regulation and has been implicated in several developmental processes. In this study, we describe the cloning and expression patterns of five Brunol genes in Xenopus laevis. Among them, only Brunol2 is maternally expressed and the zygotic expression of the other four Brunol genes starts at different developmental stages. During Xenopus development, Brunol1, 4-5 are exclusively expressed in the nervous system including domains in the brain, spinal cord, optic and otic vesicles. Brunol2 and 3 are expressed in both the somatic mesoderm and the nervous system. Brunol2 is also extensively expressed in the lens. In transfected Hela cells, BRUNOL1, 2 and 3 proteins are localized in both the cytoplasm and the nucleus, while BRUNOL4 and 5 are only present in the cytoplasm, indicating their different functions.

采用心脏灌流法去除组织内红细胞制备分散的细胞悬液的方法

本发明涉及一种采用心脏灌流法去除组织内红细胞制备分散细胞悬液的方法，属生物医学领域。本发明与公知方法不同之处在于用心脏灌流分离组织中的红细胞。本发明具有方法简便快捷，去除组织中的红细胞效果佳，分散的组织细胞的存活率高等优点，可用于制备各种组织的分散的细胞悬液。

普罗托品在制备治疗老年性痴呆症药物中的应用

本发明属生物医学技术领域，具体地说，涉及普罗托品在制备治疗老年性痴呆症药物中的应用。本发明选用从滇产植物紫金龙中用现有技术提取的纯天然产物普罗托品，通过用普罗托品对小鼠记忆获得障碍、胆碱酯酶、小鼠神经细胞钠、钾和钙离子通道的影响的研究，表明普罗托品具有胆碱酯酶抑制作用和非选择性离子通道阻断作用，即具有胆碱酯酶抑制剂、吡咯烷酮类化合物和银杏叶提取物的优点，对老年性痴呆症等各种记忆障碍会有更好的治疗作用。普罗托品具有作为制备治疗老年性痴呆症等各种记忆障碍药物的用途。

一种诱导恶性肿瘤细胞凋亡的方法和用途

本发明涉及一种诱导恶性肿瘤细胞凋亡的方法和用途，属生物医药技术领域。本发明直接用CD176(Thomsen-Friedenreich肿瘤相关抗原，TF， Galβ1-3GalNAcα1-R和Galβ1-3GalNAcβ1-R)的抗体(自然抗体、合成的人源化抗体)和配体[合成的结合CD176(TF)的重组蛋白和多肽、提取的结合CD176 (TF)的植物或动物凝集素]，结合恶性肿瘤细胞膜上的CD176(TF)，诱导恶性肿瘤细胞凋亡。本发明的诱导恶性肿瘤细胞凋亡的方法可作为恶性肿瘤生物治疗。本发明与现有技术相比，具有安全和适用范围广等优点。