Use of the STR loci D18S53, D18S59, and D18S488 in the diagnosis of Edwards’ syndrome

The aim of this study was to investigate the feasibility of using short tandem repeats (STRs) to diagnose Edwards’ syndrome (ES). Quantitative fluorescence polymerase chain reaction (QF-PCR) was performed to amplify STR loci on chromosome 18, specifically D18S53, D18S59, and D18S488. The amplified products were subjected to a fluorescence signal analysis and their application to ES diagnosis was examined. Among the 807 cases that showed normal results in the karyotype analysis, 793 showed one or two fluorescence bands with a fluorescence intensity ratio of 1:1, and 14 cases showed 3 bands, which were false-positive results. ES was diagnosed in 9 samples. The sensitivities of D18S53, D18S59, and D18S488 for the diagnosis of ES were 77.78, 44.44, and 55.56 % and the specificities were 96.16, 96.03, and 96.28 %, respectively. The combined sensitivity of the three loci for diagnosing DS was 100 % (9/9), with a specificity of 98.27 % (793/807). QF-PCR amplification of STR loci had high sensitivity, strong specificity, and was simple and rapid. Thus, it might have wide clinical applications, and could be an ideal tool for large-scale genetic and prenatal diagnosis of ES.     

西北汉族人群D10S1248、D2S441、D1S1677、D9S1122、D10S1435等5个miniSTR基因座的遗传多态性及其法医学应用

为了调查D10S1248、D2S441、D1S1677、D9S1122、D10S1435等5个miniSTR(mini short tandem repeats)基因座在西北汉族人群中的遗传多态性、遗传稳定性及其在陈旧降解检材中的法医学应用价值, 文章采用荧光PCR和基因分型技术对西北汉族154份无关个体血样、10个家系血液样本及10份陈旧降解检材进行片段长度分析。在西北汉族人群中, 5个miniSTR基因座分别检测出了8、7、7、6、7个等位基因, 等位基因频率分布符合Hardy-Weinberg平衡定律, 杂合度(Heterozygosity, H)为0.662~0.792, 个人识别率(Power of discrimination, PD)为0.869~0.915, 非父排除率(Power of exclusion, PE)为0.382~0.585, 多态信息含量(Polymorphism informa-tion content, PIC)为0.650~0.750。家系和陈旧降解检材的研究表明, 5个miniSTR基因座具有高度的遗传稳定性, 可对陈旧降解检材DNA进行有效的分型。5个miniSTR基因座适合作为西北汉族人群的遗传标记, 用于陈旧降解检材的法医学个人识别和亲权鉴定案件中。     

A polymorphism of the rat T-cell receptor β-chain variable gene 13 (BV13S1) correlates with the frequency of BV13S1-positive CD4 cells

Three rat BV13S1 alleles (T-cell receptor β-chain variable gene 13) were characterized by new BV13S1-allele specific monoclonal antibodies (18B1 and 17D5) and sequence analysis of expressed and genomic BV13S1. Two alleles were functional and designated BV13S1A1 present in strains LEW, BUF, PVG, and BV13S1A2 present in BN and WF. Their products differed by six amino acids, two of them in complementarity-determing region (CDR)1 and one in CDR2. A third nonfunctional allele, BV13S1A3P, was found in strains F344 and DA. Apart from a single nucleotide insertion, it was identical to BV13S1A2. All 12 rat strains tested showed association of TCRBC1 with BV8S2/4 alleles but not with the BV13S1 alleles, which may reflect a different gene order of the rat BV compared to mouse. BV13S1A1-encoded T-cell receptors (TCRs) which bind both monoclonal antibody (mAb) 18B1 and mAb 17D5 are over-represented in the CD4 lymphocyte subset. BV13S1A2-encoded TCRs which are stained by mAb 18B1 but not by mAb 17D5 show a slight CD8-biased expression. Preferential usage of BV13S1A1-positive TCRs by CD4 but not by CD8 cells in (LEW×WF)F1 hybrids and cosegregation of BV13SA1 and increased frequency of BV13S1 TCR-positive CD4 cells in a (LEW×BN)×BN backcross suggest structural differences of the two allelic products as the reason for their contrasting CD4/CD8 subset bias. Received: 6 October 1999 / Revised: 25 November 1999     

人体基因组D14S13位点限制性片段长度多态性的研究

应用pCE1.2探针,分析了296个无关个体基因组D14S13位点Haelll酶解的限制性片段长度多态性（RFLP）,进行了等位基因频率的分布统计,发现至少含有66个等位基因,位点杂合性大于93％,在对8个家系93名相关个体的分析中,没有发现突变发生,此RFLP分析方法已初步应用于法医学的个人识别和亲权鉴定中,并对准确度进行了计算。》     

2D relayed anisotropy correlation NMR: Characterization of the 13C′ chemical shift tensor orientation in the peptide plane of the dipeptide AibAib

An approach to the determination of the orientation of the carbonyl chemical shift (CS) tensor in a ¹³C-¹⁵N-¹H dipolar coupled spin network is proposed. The method involves the measurement of the Euler angles of the ¹³C-¹⁵N and ¹⁵N-¹H dipolar vectors in the ¹³C CS tensor principal axes system, respectively, via a ¹³C-¹⁵N REDOR experiment and by a 2D relayed anisotropy correlation of the ¹³C CSA (₂) and ¹⁵N-¹H dipolar interaction (₁). Via numerical simulations the sensitivity of the ₁ cross sections of the 2D spectrum to the Euler angles of the ¹⁵N-¹H bond vector in the ¹³C CSA frame is shown. Employing the procedure outlined in this work, we have determined the orientation of the ¹³C CS tensor in the peptide plane of the dipeptide AibAib-NH₂ (Aib = -aminoisobutyric acid). The Euler angles are found to be (_CN, _CN) = (34° ± 2°, 88° ± 2° ) and (_NH, _NH) = (90° ± 10°, 80° ± 10° ). From the measured Euler angles it is seen that the ₃₃ and ₂₂ components of the ¹³C CS tensor approximately lie in the peptide plane.     

N13D、S40E点突变提高木聚糖酶XYNB的热稳定性

对来源于Streptomyces olivaceoviridis的高比活木聚糖酶XYNB进行同源建模和序列比较,设计了N13D、S40E的定点突变,以期改善中温酶XYNB的热稳定性。突变酶N13D、S40E分别在毕赤酵母中表达,经纯化后与野生型酶XYNB(同样经毕赤酵母表达后纯化)进行酶学性质比较,结果表明,突变酶N13D和S40E在70℃处理5min,热稳定性比XYNB分别提高了24.76%和14.46%;突变酶N13D的比活性比XYNB提高了22%。在其他性质方面突变酶N13D、S40E与野生型酶XYNB基本相似。通过对木聚糖酶XYNB的定点突变,提高了该酶的热稳定性,并为结构与功能的进一步研究提供了材料。     

Characterization and Genetic Study of the Ovine α S2 -Casein (CSN1S2) Allele B

An interesting and quite complex protein pattern has been described at ovine milk proteins but the genetic control of the variation observed was assessed only in few cases. The aim of this work was to characterize the ovine α _s2 -casein (CSN1S2) B variant, first observed in the Italian Gentile di Puglia, a fine-wooled ovine breed, and to investigate its occurrence in two further breeds, the Sarda and Camosciata, which are the most widespread dairy breeds in Italy. The B variant differs from the most common form A with two amino acid exchanges: Asp₇₅ → Tyr₇₅ and Ile₁₀₅ → Val₁₀₅. The first substitution, resulting in a loss of a negative charge, is responsible for the higher isoelectric point of the B protein variant, which allows its detection by isoelectric focusing electrophoresis (IEF). The occurrence of CSN1S2*B in Sarda and Comisana was demonstrated. Since the Asp₇₅ → Tyr₇₅ substitution modifies the protein electric charge, milk properties may result affected to some extent.     

Characterization of the S289A,D mutants of yeast cystathionine β-synthase

Cystathionine β-synthase (CBS) catalyzes the pyridoxal 5′-phosphate (PLP)-dependent condensation of l-serine and l-homocysteine to form l-cystathionine in the first step of the reverse transsulfuration pathway. Residue S289 of yeast CBS, predicted to form a hydrogen bond with the pyridine nitrogen of the PLP cofactor, was mutated to alanine and aspartate. The k_cat/K_m^l-Ser of the S289A mutant is reduced by a factor of ～ 800 and the β-replacement activity of the S289D mutant is undetectable. Fluorescence energy transfer between tryptophan residue(s) of the enzyme and the PLP cofactor, observed in the wild-type enzyme and diminished in the S289A mutant, is absent in S289D. These results demonstrate that residue S289 is essential in maintaining the properties and orientation of the pyridine ring of the PLP cofactor. The reduction in activity of ytCBS-S289A suggests that ytCBS catalyzes the α,β-elimination of l-Ser via an E1cB mechanism.     

p34cdc2: the S and M kinase?

In the yeast cell cycle, the induction of two very different processes, DNA synthesis (S-phase) and mitosis (M-phase), requires the same serine/threonine-specific protein kinase p34cdc2, which has been highly conserved through evolution. On the basis of work conducted largely in multicellular eukaryotes, it has recently been suggested that p34cdc2 is able to perform these two mutually exclusive roles by phosphorylating different sets of substrates through a cell cycle-dependent association with other proteins that dictate the substrate specificity of the protein kinase. To recognize its mitotic substrates, p34cdc2 associates with one of the cyclins--a family of proteins of two distinct but related types (A and B) characterized by their periodic destruction at each mitosis. In interphase, the formation of a complex between p34cdc2 and another protein (or proteins) would allow the phosphorylation of a different set of proteins involved in the G1 to S transition. This review focuses on the evidence for this appealing simple model and the nature of the putative substrates proposed.     

A QM protein–ligand investigation of antipsychotic drugs with the dopamine D2 Receptor (D2R)

The dopamine D2 Receptor (D2R) is a member of the G-Protein-Coupled Receptor family and plays a critical role in neurotransmission activities in the human brain. Dysfunction in dopamine receptor signaling may lead to mental health illnesses such as schizophrenia and Parkinson’s disease. D2R is the target protein of the commonly used antipsychotic drugs such as risperidone, clozapine, aripiprazole, olanzapine, ziprasidone, and quetiapine. Due to their significant side effects and non-selective profiles, the discovery of novel drugs has become a challenge for researchers working in this field. Recently, our group has focused on the interactions of these drug molecules in the active site of the D2R using different in silico approaches. We here compare the performances of different approaches in estimating the drug binding affinities using quantum chemical approaches. Conformations of drug molecules (ligands) at the binding site of the D2R taken from the preliminary docking studies and molecular dynamics simulations were used to generate protein–ligand interaction models. In a first approach, the BSSE-corrected interaction energies of the ligands with the most critical amino acid Asp114 and with the other amino acids closest to ligands in the binding cavity were calculated separately by density functional theory method in implicit water environment at the M06-2X/6-31 g(d,p) level of the theory. In a second approach, ligand binding affinities were calculated by taking into consideration not only the interaction energies but also deformation and desolvation energies of ligands with surrounding amino acid residues, in a radius of 5 Å of the protein-bound ligand. The quantum mechanically obtained results were compared with the experimentally obtained binding affinity values. We concluded that although H-bond interactions of ligands with Asp114 are the most dominant interaction in the binding site, if van der Waals and steric interactions of ligands which have cumulative effect on the ligand binding are not included in the calculations, the interaction energies are overestimated.     

Does the 13C of foliage‐respired CO2 and biochemical pools reflect the 13C of recently assimilated carbon?

Isotopic labelling experiments were conducted to assess relationships among ¹³C of recently assimilated carbon ( δC _A), foliage respiration ( δC _F), soluble carbohydrate ( δC _SC), leaf waxes ( δC _LW) and bulk organic matter ( δC _OM). Slash pine, sweetgum and maize were grown under ¹³C depleted CO₂ to label biomass and then placed under ambient conditions to monitor the loss of label. In pine and sweetgum, δC _F of labelled plants (∼−44 and −35‰, respectively) rapidly approached control values but remained depleted by ∼4–6‰ after 3–4 months. For these tree species, no or minimal label was lost from δC _SC, δC _LW and δC _OM during the observation periods. δC _F and δC _SC of labelled maize plants rapidly changed and were indistinguishable from controls after 1 month, while δC _LW and δC _OM more slowly approached control values and remained depleted by 2–6‰. Changes in δC _F in slash pine and sweetgum fit a two-pool exponential model, with the fast turnover metabolic pool (∼3–4 d half-life) constituting only 1–2% of the total. In maize, change in δC _F fits a single pool model with a half-life of 6.4 d. The ¹³C of foliage respiration and biochemical pools reflect temporally integrated values of δC _A, with change in isotopic composition dampened by the size of metabolic carbon reserves and turnover rates.     

广州汉族人群D12S391和D11S554基因座序列变异

为研究高度变异的sTR基因座核心序列结构,对广州汉族人群突变率较高的D12S391和D11S554基因座等位基因进行了序列分析。结果显示D12S391基因座核心序列结构为(AGAT)8～17(AGAC)6～10(AGAT)0～1,其较小片段等位基因(15～18)仅表现为第1个重复单位(AGAT)数目的变异,而较大片段等位基因(19～27),可表现为第2或第3个重复单位数目的变异。发现有4种新的等位基因,分别命名为22″、23″、24′″和27。D11S554基因座核心序列结构更复杂,有5种核心序列,其中3种具有相同的基本结构(AAAGG)(AAAG)4(AAAGG)2～3,(AAAG)13～19。其大片段等位基因(219～249)核心序列结构中,既有四核苷酸重复,还有五核苷酸重复,以及单个硷基的变异、硷基插入或缺失。两基因座均存在序列异质性。结果表明D12S391和D11S554基因座属复杂重复类型,为其准确分型增加了难度,首先需建立相应群体的等位基因分型标准物。     

D3S1358、D21S11和FGA复合扩增系统的研究

目的：研究D3S1358、D21S11和FGA3个STR基因座复合扩增的最佳体系。方法：设计正交实验优化复合扩增最佳反应条件,然后采用聚丙烯酰胺凝胶电泳分离显带技术检测扩增片段。结果：获得复合扩增条件各反应因素较为满意的参数。     

北京地区汉族群体D1S1612和D18S535基因座的遗传多态性

采用扩增片段长度多态性（Amp-FLP）分型技术,调查中国北京地区汉族群体D1S1612、D18S535 基因座的遗传多态性,获得等位基因频率分布。结果显示, D1S1612检出9个等位基因,25种基因型, D18S535检出9个等位基因,27种基因型。两个STR基因座的杂和度（H）分别为0.779、0.887;个人识别率（Dp）分别为0.901、0.927;非父排除率（PE）分别为0.564、0.770;多态信息容量（PIC）分别为0.723、0.796,卡方检验表明两个STR 基因座基因型频率分布符合Hardy-Weinberg平衡 (P>0.01 )。D1S1612和D18S535 基因座均属高杂合度、高识别能力的遗传标记,可用于法庭科学亲子鉴定和个人识别。 Abstract: To investigate the genetic polymorphism of D1S1612 and D18S535 in Han population of Beijing. Amp-FLP method was used. 9 alleles, 25 genotypes were observed for D1S1612 locus; and 9 alleles and 27 genotypes for D18S535 locus. All allele frequencies, heterozygosity (H), discrimination power (Dp), exclusion of paternity probability (PE) and polymorphism information content (PIC) were calculated. The allele distributions of the two loci were conformed to Hardy-Weinberg equilibrium (P>0.01). According to the results obtained in this study, it is suggested that both D1S1612 and D18S535 are useful genetic markers for individual identification and paternity testing in forensic science practice as well for genetic study.     

Elite swimmers and the D allele of the ACE I/D polymorphism

A polymorphism of the human angiotensin-1-converting enzyme (ACE) gene has been identified in which the presence (insertion, I allele) of a 287-bp fragment rather than the absence (deletion, D allele) is associated with lower ACE activity. Several recent studies have shown an association of the I allele with endurance performance, it being found with excess frequency in elite distance runners, rowers and mountaineers. Other workers using heterogeneous cohorts of athletes from mixed sporting disciplines have found no such association. An increasing linear trend of I allele frequency with the distance run amongst Olympic runners and an excess of the D allele amongst sprinters led us to examine whether the ratio of I and D alleles in swimmers competing over different distances would also vary. Swimmers (n=120) from the European and Commonwealth championships and an American college team had their ACE genotype determined and their gene and allele frequencies compared with several control groups, the most closely age-matched of which were 1,248 military recruits. Of the 103 Caucasians, there was a significant excess of the D allele compared with this control group only in the truly elite swimmers of the European and Commonwealth championships (P=0.004). This association remained in those competing over shorter distances (P=0.005 for 400 m and below) but not in the longer events. These findings were confirmed in three further large control groups. A population association study testing whether a genetic marker (the ACE I/D polymorphism) occurs more frequently in cases (elite athletes) than in controls therefore requires a homogeneous cohort of subjects from the same sporting discipline.     

基于26S rDNA D1/D2序列分析酱香型白酒酒醅中酵母菌的群落结构

【目的】比较分析不同发酵阶段的酱香型白酒北大仓酒醅中酵母菌的群落结构。【方法】酒醅样本采集自黑龙江省北大仓白酒不同阶段的发酵窖池,采用平板分离技术获得大量酵母菌纯培养物;基于26S rDNA D1/D2区域的碱基序列,准确鉴定酒醅中的酵母菌;根据酵母菌的数量变化、种类组成、相对频率、多样性指数及其相似性系数,研究酱香型北大仓白酒酒醅中酵母菌的群落结构特征。【结果】北大仓白酒酒醅中酵母菌数量在酿造过程中的变化,遵循着"升高-降低-升高-降低"的规律;第1轮"升高-降低"的过程表现出急剧的"速升速降"特点,而第2轮"升高-降低"的过程则具有相对来说比较温和的"缓升缓降"特征。酒醅内的酵母菌鉴定为8属13种,总体多样性指数处于较高的水平,发酵初期酵母菌多样性指数逐渐增加,最高的多样性指数(1.89)出现在发酵第5 d的酒醅内;然后又逐渐下降,发酵至第11 d时降至最低(0.66);酒醅发酵至13 d时开始直至发酵末期,多样性指数呈现出波动性变化。北大仓白酒酒醅中酵母菌的种类组成总体来说比较接近,大多数情况下相似性系数都高于0.5,说明在白酒酿造过程中酵母菌的组成和分布处于一个相对稳定的状态。对于北大仓白酒酒醅内存在的常见酵母菌来说,Issatchenkia orientalis、Pichia kudriavzevii和Saccharomyces cerevisiae可能在酿造过程中发挥更重要的作用。【结论】北大仓等酱香型白酒的酒醅内存在着丰富的酵母菌资源,阐明酒醅内酵母菌的群落结构可以为解析酱香型白酒的发酵机理提供基础数据和理论基础。     

基于核糖体ITS区和28S rRNA D2～D3区的短体线虫系统发育研究

短体线虫又称根腐线虫,是世界分布最为广泛和最具破坏性的迁徙性植物内寄生线虫之一.本研究根据214条核糖体ITS序列,218条核糖体28S大亚基D2～D3序列,应用MEGA 4.0软件,通过邻接法(NJ)构建了短体线虫的系统发育树.结果发现,2个系统树在整体上大致相似,仅在小的分支上存在差异.基于ITS序列的系统树将25种短体线虫至少分为8组,相应的基于D2～D3序列的系统树将23种短体线虫至少分为7组.其中,有3个大组内部的系统发育关系比较清晰.根据本研究的系统发育分析仍然无法从总体上确定短体线虫种间的系统发育关系.     

Penam Sulfones and β-Lactamase Inhibition: SA2-13 and the Importance of the C2 Side Chain Length and Composition

β-Lactamases are the major reason β-lactam resistance is seen in Gram-negative bacteria. To combat this resistance mechanism, β-lactamase inhibitors are currently being developed. Presently, there are only three that are in clinical use (clavulanate, sulbactam and tazobactam). In order to address this important medical need, we explored a new inhibition strategy that takes advantage of a long-lived inhibitory trans-enamine intermediate. SA2-13 was previously synthesized and shown to have a lower k _react than tazobactam. We investigated here the importance of the carboxyl linker length and composition by synthesizing three analogs of SA2-13 (PSR-4-157, PSR-4-155, and PSR-3-226). All SA2-13 analogs yielded higher turnover numbers and k _react compared to SA2-13. We next demonstrated using protein crystallography that increasing the linker length by one carbon allowed for better capture of a trans-enamine intermediate; in contrast, this trans-enamine intermediate did not occur when the C2 linker length was decreased by one carbon. If the linker was altered by both shortening it and changing the carboxyl moiety into a neutral amide moiety, the stable trans-enamine intermediate in wt SHV-1 did not form; this intermediate could only be observed when a deacylation deficient E166A variant was studied. We subsequently studied SA2-13 against a relatively recently discovered inhibitor-resistant (IR) variant of SHV-1, SHV K234R. Despite the alteration in the mechanism of resistance due to the K→R change in this variant, SA2-13 was effective at inhibiting this IR enzyme and formed a trans-enamine inhibitory intermediate similar to the intermediate seen in the wt SHV-1 structure. Taken together, our data reveals that the C2 side chain linker length and composition profoundly affect the formation of the trans-enamine intermediate of penam sulfones. We also show that the design of SA2-13 derivatives offers promise against IR SHV β-lactamases that possess the K234R substitution.