大肠癌中p53基因突变的研究

应用聚合酶链反应(PCR)──单链构型多态性(SSCP)结合银染法对14例大肠癌p53基因的第4、第5─6和第7外显子进行了点突变的研究,结果共检测出6例点突变,而且发现各外显子的突变频率存在差异。另外,利用购自ATCC的两个探针 (p53cDNA探针和pYNZ22探针)对大肠癌中p53基因的杂合性失去进行了研究,在14例大肠癌中共检出6例杂合性丢失。将点突变检测结果同杂合性丢失结果进行比较分析, 并着重探讨了大肠癌中p53基因失活导致肿瘤的作用方式。 Abstract:The exons 4-7 of p53 gene were examined in 14 colorectal Cancer patients by using PCR-SSCP-silver staining method.The results showed 6 cases of point mutation and the mutation frequencies of exons were different from each other.p53 cDNA and pYNZ22 VNTR were used as probes to examine LOH(Loss of heterozygosity)of 14 colorectal cancers.6 cases with LOH were found.The results of present research suggest that mutation and LOH of p53 gene are critical events in the progress and development of Cancer.There were different kinds of inactivation model of p53 gene in the process of development of cancer and transformation of cells.     

Mechanism of MAT alpha donor preference during mating-type switching of Saccharomyces cerevisiae.

During homothallic switching of the mating-type (MAT) gene in Saccharomyces cerevisiae, a- or alpha-specific sequences are replaced by opposite mating-type sequences copied from one of two silent donor loci, HML alpha or HMRa. The two donors lie at opposite ends of chromosome III, approximately 190 and 90 kb, respectively, from MAT. MAT alpha cells preferentially recombine with HMR, while MATa cells select HML. The mechanisms of donor selection are different for the two mating types. MATa cells, deleted for the preferred HML gene, efficiently use HMR as a donor. However, in MAT alpha cells, HML is not an efficient donor when HMR is deleted; consequently, approximately one-third of HO HML alpha MAT alpha hmr delta cells die because they fail to repair the HO endonuclease-induced double-strand break at MAT. MAT alpha donor preference depends not on the sequence differences between HML and HMR or their surrounding regions but on their chromosomal locations. Cloned HMR donors placed at three other locations to the left of MAT, on either side of the centromere, all fail to act as efficient donors. When the donor is placed 37 kb to the left of MAT, its proximity overcomes normal donor preference, but this position is again inefficiently used when additional DNA is inserted in between the donor and MAT to increase the distance to 62 kb. Donors placed to the right of MAT are efficiently recruited, and in fact a donor situated 16 kb proximal to HMR is used in preference to HMR. The cis-acting chromosomal determinants of MAT alpha preference are not influenced by the chromosomal orientation of MAT or by sequences as far as 6 kb from HMR. These data argue that there is an alpha-specific mechanism to inhibit the use of donors to the left of MAT alpha, causing the cell to recombine most often with donors to the right of MAT alpha.     

Effects of anti-peptide antibodies against the second extracellular loop of human M2 muscarinic acetylcholine receptors on transmembrane potentials and currents in guinea pig ventricular myocytes

The effects of anti-peptide antibodies against the second extracellular loop of human M2 muscarinic receptor on transmembrane potentials and currents in guinea pig single ventricular cells were analyzed using whole-cell patch clamp technique. These effects were compared with those of the muscarinic receptor agonists carbachol and acetylcholine. The antibodies shortened the action potential duration in a dose-dependent manner. By using a ramp or step rectangular pulse protocol, it was found that the antibodies increased the outward K⁺ current and decreased the inward basal I Ca significantly. The reversal potential of both carbachol-and antibody-induced extra currents were close to –80 mV, being in proximity to the calculated E_k of –90 mV. A -adrenergic receptor agonist, isoprenaline, prolonged the action potential and increased the overshoot which could be inhibited by both antibody and carbachol. Isoprenaline increased inward I_ca and outward I_k simultaneously. Both antibody and carbachol could significantly reduce the isoprenaline-stimulated I_Ca but not the isoprenaline-stimulated I_k. The antibody- or carbachol-induced outward K⁺ current and the depressant effects of antibody and carbachol on isoprenaline-stimulated I_ca were partially antagonized by atropine. These results suggest that the anti-M2 muscarinic receptor antibodies display a stimulatory activity similar to muscarinic receptor agonist on the receptor-mediated electrophysiological events.     

Fluorescence and Fourier-transform infrared spectroscopic studies on the role of disulfide bond in the calcium binding in the 33 kDa protein of Photosystem II

The 33 kDa protein of Photosystem II has one intrachain disulfide bond. Fluorescence spectroscopy shows that the major groups in the protein that bind to Ca²⁺ should be the carboxylic side groups of glutamic acid and/or aspartic acid. Fluorescence and Fourier-transform infrared (FTIR) spectroscopic studies indicate that the conformation of the 33 kDa protein is altered upon reduction, while the reduced protein still retains the secondary structure. FTIR spectroscopy also shows that the metal ions induce a relative decrease of unordered structure and -sheet, and a substantial increase of -helix in both the intact and the reduced 33 kDa protein. This indicates that the addition of cations results in a much more compact structure and that both the intact and the reduced 33 kDa proteins have the ability to bind calcium. The above results may suggest that the disulfide bridge is not essential for calcium binding.Abbreviations CD circular dichroism - FTIR Fourier transform infrared - La lanthanum - PS photosystem - Tb terbium     

Site-directed mutagenesis of the CP 47 protein of photosystem II: 167W in the lumenally exposed loop C is required for photosystem II assembly and stability

The intrinsic chlorophyll-protein CP 47 is a component of photosystem II which functions in both light-harvesting and oxygen evolution. Using site-directed mutagenesis we have produced the mutant W167S which lies in loop C of CP 47. This strain exhibited a 75% loss in oxygen evolution activity and grew extremely slowly in the absence of glucose. Examination of normalized oxygen evolution traces indicated that the mutant was susceptible to photoinactivation. Analysis of the variable fluorescence yield indicated that the mutant accumulated very few functional PS II reaction centers. This was confirmed by immunoblotting experiments. Interestingly, when W167S was grown in the presence of 20 M DCMU, the mutant continued to exhibit these defects. These results indicate that tryptophan 167 in loop C of CP 47 is important for the assembly and stability of the PS II reaction center.     

Structure and analysis of phospholipase D gene from Ricinus communis L

Phospholipase D (PLD; EC 3.1.4.4) has been proposed to play a pivotal role in various cellular processes, but molecular understanding of this enzyme is rather limited. This report describes the nucleotide sequence, structure, and genomic organization of a PLD gene from castor bean (Ricinus communis L. cv. Hale). The PLD gene was isolated from a castor bean genomic library using the PLD cDNA as a hybridization probe. Sequence comparison with the PLD cDNA revealed that the PLD gene consisted of four exons and three introns, one of which interrupts the 5-untranslated region. Southern blot analysis indicated that the cloned PLD gene was present as a single-copy gene, and yet there were other PLD or PLD-related sequences in the castor bean genome.     

懒猴属的核糖体DNA变异及其种间分化关系

用１５种限制性内切酶和人２８Ｓ、１８ＳｒＤＮＡ探针构建了懒猴属各物种核糖体ＤＮＡ重复单位的限制性内切酶图谱。在进化速率较高的非转录间隔区,在大、中、小懒猴中分别定位了２３、２４、２４个酶切位点。大懒猴与中懒猴有１２个位点不同,与小懒猴有１４个位点不同,而中、小懒猴间则只有一个位点的差异。经过计算,大懒猴与中懒猴的遗传距离值为１２．６５％,与小懒猴的差异为１４．２４％,中、小懒猴间的差异则仅为０．７     

云南姬鼠的蛋白多态性及其遗传分化关系

本文采用蛋白电泳技术对来源于云南省若干地区的姬鼠属（Ａｐｏｄｅｍｕｓ）的３种姬鼠──高山姬鼠（Ａ．ｃｈｅｖｒｉｅｒｉ）８只,中华姬鼠（Ａ．ｄｒａｃｏ）３只和大耳姬鼠（Ａ．ｌａｔｒｏｎｕｍ）１只,以及作为外群的同科的绒鼠属的大绒鼠（Ｈａｐａｌｏｍｙｓｄｅｌａｌｏｒｉ）３只进行了分析。共检测遗传座位２７个,发现２１个座位存在多态性。根据蛋白多态的数据对研究对象进行遗传分化关系的探讨,用系统分析软件ＰＨＹＬＩＰ计算它们之间的分化关系,得到了一棵无根系统树。结果表明,作为外群的大绒鼠明显不同于其它３种姬鼠而聚在最外面。８只高山姬鼠个体汇聚成独立的一支,中华姬鼠的３个个体也聚成一支,但大耳姬鼠却聚在中华姬鼠一支中,因此我们认为大耳姬鼠同中华姬鼠的分化时间可能比较晚近。     

藏酋猴的分类与分布

本文基于外部形态,毛色,头骨特征和地理分布对藏酋猴进行了分类整理,认为藏酋猴在不同地理区域之间的差异已达到了亚种水平,可分为４个亚种（包括两新亚种）：Ｍ．ｔｈｉｂｅｔａｎａｔｈｉｂｅｔｎａ,Ｍ．ｔｈｉｂｅｔａｎａｐｕｌｌｕｓ．Ｍ．ｔｈｉｂｅｔａｎａｈｕａｎｇｓｈａｎｅｎｓｉｓｓｕｂｓｐ．ｎｏｖ．和Ｍ．ｔｈｉｂｅｔａｎａｇｕｉｚｈｏｕｅｎｓｉｓｓｕｂｓｐ．ｎｏｖ。