DNA生物传感器进展

近几年来,酶传感器、免疫传感器及微生物传感器等发展较为成熟,而DNA生物传感器的研究相对较少.文章从核酸杂交的原理出发介绍了DNA生物传感器的工作原理,举例说明了电化学、光学和声学等几种典型的DNA生物传感器,指出了其固有的优缺点,肯定了DNA传感器发展前景.     

激活淋巴细胞cDNA单链库的构建

介绍一种激活淋巴细胞cDNA单链库构建方法,此库具有良好的模板活性. 用PCR法从此库中克隆了多种中国人源性的细胞因子.     

藻胆蛋白复合物的合成及其分子内能量传递

通过偶联剂3-(2-吡啶联巯基)丙酸N-羟基琥珀亚胺酯(SPDP)及改变配料比, 合成了两个R-藻红蛋白(R-PE)与C-藻蓝蛋白(C-PC)的复合物A和B.利用吸收光谱确定了分子内R-PE与C-PC的摩尔比为6∶1和2∶1. 通过荧光光谱, 观察到能量传递现象, 并计算出能量传递效率为63%和88%.证明分子内能量传递效率很高. 二硫苏糖醇(DTT)还原连接R-PE与C-PC的二硫桥键后, 能量传递被阻断. 这一现象进一步证明复合物中存在分子内能量传递.     

博莱霉素-Ce(Ⅲ)对DNA的切断机理初探

根据外切核酸酶Ⅲ酶解博莱霉素-Ce(Ⅲ)［BLMA5-Ce(Ⅲ)］作用过的双链直线型DNA时, 酶解速率明显增大, 酶解产物除5′-dAMP、5′-dGMP、5′-dCMP和5′-dTMP 4种单核苷酸外, 还有其他成分存在的实验事实, 推测出BLMA5-Ce(Ⅲ)在DNA双链的特定部位沿5′→3′的方向切断磷酸二酯键, 使DNA的双链上形成多个暴露的3′-OH末端.     

冬虫夏草(Cordyceps sinensis)的随机扩增多态DNA及其遗传分化

本文对来自青藏高原3个区域5个具有代表性地方的13个冬虫夏草(Cordyceps sinensis[Berk.] Sacc.)样本进行随机扩增多态DNA(RAPD)分析。19个随机引物获得的RAPD谱带清晰并呈现多态,单个引物获得的RAPD片段数在3～10个之间。该19个引物在每个样本中扩增的RAPD片段总数平均约为65个。基于遗传距离分析,受试的13个冬虫夏草样本中,来自同一地方的样本间遗传差异甚微,同一区域不同地方的样本间遗传差异较大,不同区域的样本间遗传差异最大。这说明冬虫夏草地理群体间存在着遗传分化。应用UPGMA和NJ方法构建的分子系统树显示,来自5个地方的冬虫夏草实际上可以归并为显著不同的3个组,对应于样本来源的3个区域,提示RAPD标记在冬虫夏草群体中有显著的地区特异性。我们的结果还表明,冬虫夏草地理群体间的遗传差异度与地理距离呈正相关。因此,RAPD作为有效的遗传标记,可用于研究冬虫夏草的遗传多样性、起源以及系统演化等。     

利用染色体涂染方法建立人和白眉长臂猿(Hylobates hoolock)的比较染色体图谱

除人Y染色体外,本文采用生物素标记的人全部整条染色体特异探针与白眉长臂猿(Hylobates hoolock)有丝分裂中期分裂相进行染色体原位杂交即染色体涂染法以研究人和白眉长臂猿染色体之间的同源性。在白眉长臂猿18对常染色体上检测出了与人22对常染色体同源的59对染色体片段,确定了人和白眉长臂猿之间的精度较高的染色体连锁群。结果表明:自人与白眉长臂猿的祖先分歧以来,大量的染色体间重排(至少发生了39次易位)和染色体内的重排导致了二者核型的差异。根据杂交结果绘制了首份人和白眉长臂猿比较染色体图谱,并结合已有的人和白掌长臂猿(Hylobates lar)(2n=44)和合趾长臂猿(Hylobates syndactylus)(2n=50)的比较染色体图谱对长臂猿属的染色体进化作了初步的探讨。     

Ganglioside GM3 modulates conformation of reconstituted Ca~(2 ) -ATPase

Using steady-state fluorescence and nanosecond time-resolved fluorescence techniques, the Ca 2 -ATPase conformational changes induced by ganglioside GM3 were studied with different quenchers. The results showed that GM3 could significantly increase the lifetime of intrinsic fluorescence of Ca2 -ATPase reconstituted into proteoliposomes, and could also weaken the intrinsic fluorescence quenching by KI or hypocrellin B, HB. Further-more, by using quenching kinetic analysis of the time-resolved fluorescence, in the presence of GM3, the quenching constant (Ksv) and quenching efficiency were significantly lowered. The obtained results suggest that the oligosaccha-ride chain and the ceramide moieties of the GM3 molecule could interact with its counterparts of the Ca2 -ATPase re-spectively, thus change the conformation of the hydrophobic domain of the enzyme, making the tryptophan residues in different regions shift towards the hydrophilic-hydrophobic interface, and hence shorten the distance between the hy     

Double-gating mechanism and diversity of an adenosine triphosphate (ATP)-sensitive K~ channel in neurons acutely dissociated from rat neocortex

Classically, ion channels are classified into 2 groups: chemical-sensitive (ligand-gated) and voltage-sensitive channels. Single ATP-sensitive K (K-ATP) channel currents were recorded in acutely dissociated rat neo-cortical neurons using patch clamp technique. A type of K-ATP channel has been found to be gated not only by intra-cellular ATP, but also by membrane potential ( Vm) , and proved to be a novel mechanism underlying the gating of ion channels, namely bi-gating mechanism. The results also show that the K-ATP channels possess heterogeneity and di-versity. These types of K-ATP channels have been identified in 40.12% of all patches, which are different in activa-tion-threshold and voltage-sensitivity. The present experiment studied the type-3 K-ATP channel with a unitary con-ductance of about 80 pS in detail ( n = 15). Taking account of all the available data, a variety of K-ATP channels are suggested to exist in body, and one type of them is bi-gated by both chemical substances and membrane poten     

Sequence analysis of lacZ- mutations induced by ion beam irradiation in double-stranded M13mp18DNA

While M13mp18 double-stranded DNA was irradiated with ion beam, and transfected intoE. coli JM103, a decrease of transfecting activity was discovered. The lacZ^- mutation frequency at 20% survival could reach (3.6–16.8) × 10⁴, about 2, 3–10 times that of unirradiated M13DNA. Altogether, 27 IacZ^-mutants were selected, 10 of which were used for sequencing. 7 of the sequenced mutants show base changes in 250-bp region examined (the remaining 3 mutants probably have base changes outside the regions sequenced). 5 of the base-changed mutants contain more than one mutational base sites (some of them even have 5–6 mutational base sites in 250-bp region examined); this dense distribution of base changes in polysites has seldom been seen in X-rays, Y-rays or UV induced DNA mutations. Our experiments also showed that the types of base changes include transitions(50%), transversions (45%) and deletion (5%); no addition or duplication was observed. The transitions were mainly C→T and A→G; the transversions were mainly C→A and C→G. The mutations involving cytosine residue (in the template strand) constitute about 60% of all the base changes observed. In comparison with the surrounding sequences of mutational base sites, the base located between TG and CT is found to be easily substituted.     

Sequence variation and genetic diversity in the giant panda

About 336–444 bp mitochondrial D-loop region and tRNA gene were sequenced for 40 individuals of the giant panda which were collected from Mabian, Meigu, Yuexi, Baoxing, Pingwu, Qingchuan, Nanping and Baishuijiang, respectively. 9 haplotypes were found in 21 founders. The results showed that the giant panda has low genetic variations, and that there is no notable genetic isolation among geographical populations. The ancestor of the living giant panda population perhaps appeared in the late Pleistocene, and unfortunately, might have suffered bottleneck attacks. Afterwards, its genetic diversity seemed to recover to some extent. Project supported by the “8.5” Key Project of Chinese Academy of Sciences, the Chairman Foundation of Chinese Academy of Sciences, K. C. Wang Education Foundation, the Applied Basic Research Foundation of Yunnan, the National Natural Science Foundation of China, the Special Foundation for Returned Chinese Scientists, and Zoological Society of San Diego.