X染色体失活机理研究

X染色体失活是哺乳动物中为实现雌性XX个体和雄性XY个体间X染色体上基因剂量补偿作用(dosage compensation)而普遍存在的一种现象,表现为雌性个体两条X染色体中的一条结构异固缩和大范围的基因失活。由于失活基因高度甲基化,曾经认为甲基化在这一过程中发挥重要作用并据此提出一些模型,但相反的证据不断积累使人们对甲基化在这一过程中的主导作用发生怀疑。由于X     

9例罕见异常染色体新核型

我科于1985年10月一1987年10月期间 共发现9例世界首报新核型,其中从流产、早产 病人中检出斗例,闭经病人中检出3例,从阵发 性睡眠性血红蛋白尿（简称PNH）和无精症的 病人中各检出1例。现分别报告如下：     

光学基因组图谱在染色体结构异常检测中的应用

染色体结构异常（structural variations,SVs）是临床中的常见现象,主要包括染色体缺失、重复、倒位和易位等,因其会引起基因融合、断裂或拷贝数改变,常导致疾病的发生。目前临床上对于SVs的检测多采用染色体核型、芯片分析或高通量测序等细胞遗传学或常规分子遗传学技术。近年来,随着检测技术的不断发展,隐匿性染色体平衡易位和微小片段的异常在产前诊断、新生儿遗传病、血液病及肿瘤等研究领域逐渐被重视,而传统的遗传学检测方法存在较为明显的局限性。光学基因组图谱（optical genome mapping,OGM）是近年来基于全新原理开发的分子遗传学检测技术,与传统技术相比,其对常规和隐匿性SVs（平衡易位、微小片段等）都有较强的检出能力,在多领域有较高的应用价值。但是OGM也存在一定的局限性,如其因灵敏度较高而需要其他手段辅助排除假阳性结果,在近着丝粒等无标记区域无法进行检测等。本文针对传统细胞遗传学检测技术和OGM对SVs的检测能力,OGM的优势、缺陷及其在肿瘤、儿科等领域的应用进行综述。     

用间期荧光原位杂交检测卵巢癌细胞中X染色体数目的异常

为了探讨用荧光原位杂交技术（fluorescence in situ hybridization, FISH）检测卵巢癌细胞中性染色体拷贝数目异常的实验方法及其应用价值,收集18例新鲜卵巢癌组织标本,以Biotin标记的X染色体α-卫星DNA（pBamX7）探针与经处理的标本进行卵巢癌细胞核的原位杂交,分别用Avidin-FITC和Anti-avidin进行信号的检测与放大,PI复染。于Olympus AX-70型荧光显微镜下,通过WIB滤光镜观察杂交信号及其细胞核背景,并统计卵巢癌细胞核中的杂交信号颗粒数量。在显微镜下可见以Biotin标记的pBamX7探针显示绿色杂交信号,细胞核背景经PI复染显示桔红色;发现11/18（61%）卵巢癌标本中X染色体拷贝数增加,其余7例（39%）无拷贝数增加。X染色体拷贝数目增多在卵巢癌中有一定比例的发生频率,其在促进卵巢癌发病及其发展过程中起到某种作用,其意义值得进一步研究。     

染色体高层结构研究

真核细胞染色体的构筑和功能可以说是生物学中一系列没有解决的最重要的问题之一.诸如染色体中DNA序列组织,非活性染色质结构,基因活化表达时染色体结构的变化;在分化组织中活化基因的决定、维持和调控,细胞周期运行过程中染色体结构的调节,DNA复制过程中染色质的去组装和重组装机理,中期染色体结构等问题,都待于探讨研究.近年来由于分子生物学技术、生物化学技术和染色体制备技术的进展,染色体显带技术和染色体电镜技术及各物理、化学手段的应用,使得染色体的生物学研究取得了一定的进展.     

基因异常和染色体结构

DNA(脱氧核糖核酸)携带有细胞为制造蛋白质所必需的信息。一条染色体就是一个紧紧缠绕着的线状DNA分子。一条染色体的DNA伸直时其长约为5cm。正常体细胞(不是卵或精子)有23对染色体,这些染色体都具有特征性的形状。有时胚胎细胞中的染色体的形状会发生明显改变,以至能在显微镜下辨认出它们的异常。例如,染色体的一些部分会在所谓的交换过程中相互交换。交换发生在减数分裂期间,这时卵和精它     

染色体末端微小结构异常的分子细胞遗传检测

为检出易于被忽略的染色体末端微小结构异常,为生育提供指导,选取特异性7号全染色体探针,X染色体长臂探针和7q亚端粒（7q36→qter)探针,用荧光原位杂交（fluorescence in situ hybridzation,FISH)结合G显带技术分析2个病例,其中病例1有不良妊娠史并疑有末端微小易位,病例2在G显带水平已发现为X和7号染色体易位的卵巢早衰患者,结果表明,FISH确诊病例1为染色体末端的隐匿易位,病例2的易位断点得到精确定位,它不在7q36而在7q末端。应用特异性染色体探针及亚端粒探针,通过FISH技术可以确诊染色体末端区域的微小结构异常,在临床遗传学中有广泛的应用,是遗传咨询和生育指导的有效工具之一。     

47例染色体异常患者永生淋巴细胞株的建立

采用4种不同的血细胞/EB病毒（Epstein-Barr Virus,EBV）比率系列转染冻存血方法建立永生淋巴细胞株,建株成功率达97.87%,并成功地对47例染色体异常患者建立了永生淋巴细胞株,为今后不断进行的深入研究奠定了基础。     

真核细胞染色体结构研究的某些进展

生物学中的重要课题之一是真核细胞染色体结构的研究,它涉及到诸如癌变、基因调控、细胞分化、发育等重要和基础课题。目前,关于30nm 染色质纤丝如何压缩形成中期染色体仍存在分歧。其中主要有3种模型:放射环模型、螺旋模型、折叠纤维模型。特别是放射环模型最近引起广泛的注意和争论。该模型认为中期染色体是由约30nm的染色质纤丝以染色体纵轴为中心向外辐射状伸出的环集缩而成。本文拟对这一方面的研究工作进行综述和讨论。1977年,Laemmli 等以中期 Hela 细胞为材料,用硫酸葡聚糖和肝素处理,去除组蛋     

Y染色体异常29例分析

本文从1992例遗传咨询病例中收集29例Y染色体异常的病例,其中Y染色体数 目异常(47,XYY)2例;Y染色体结构异常8例:Y/Y易位1例、Yp+3例、de l(Y)3例、嵌合 体dic(Y)1例;Y染色体长度变异19例。对Y染色体这几种异常类型的遗传效应进行分析。 Abstract:Twenty nine cases of Y chromosome abnormalities were found in 1992 patients asking genetic counseling.Different kinds of Y chromosome abnormalitics were detected by G and banding techniques.These were 47,XYY(2 cascs);46,X,del(Y)(3 cascs);46,X,Yp+(3 cases);46,X,t(Y;Y)(1 case);45,X/46,X,dic(Y)(1 case) and length changes of Y chromosome(19 cases).The genetic effects of Y chromosome abnormalities have been analyzed in this report.     

Chromosomal Abnormality and Y Chromosome Microdeletion in Chinese Patients with Azoospermia or Severe Oligozoo-spermia

Chromosomal abnormality and Y chromosome microdeletion are regarded as two frequent genetic causes associated with spermatogenic failure in Caucasian population. To investigate the distribution of the two genetic defects in Chinese patients with azoospermia or severe oligozoospermia, karyotype analysis by G-banding was carried out in 358 idiopathic infertile men, including 256 patients with azoospermia and 102 patients with severe oligozoospermia, and screening of AZF region microdeletion of Y chromosome by multiplex PCR was performed in those patients without detectable chromosomal abnormality and 100 fertile controls. Of 358 patients, 39(10.9%) were found to have chromosomal abnormalities in which Klinefelters syndrome (47, XXY) was the most common chromosomal aberration. The incidence of sex chromosomal abnormality in patients with azoospermia was significantly higher than that in patients with severe oligozoospermia (12.1% vs 1%). Among the rest of the 319 patients with normal karyotype, 46 (14.4%) were found to have microdeletions in AZF region. The prevalence rates of AZF microdeletion was 15% and 13.1% in patients with azoospermia and severe oligozoospermia respectively. The microdeletion in AZFc was the most frequent deletion and all the microdeletions in AZFa were found in azoospermic patients. No microdeletion in AZF region was detected in fertile controls. In conclusion, chromosomal abnormality and AZF region microdeletion of Y chromosome might account for about 25% of Chinese infertile patients with azoospermia or severe oligozoospermia, suggesting the two abnormalities are important genetic etiology of spematogenic failure in Chinese population and it is essential to screen them during diagnosis of male infertility before in vitro assisted fertilization by introcytoplasmic sperm injection.     

中国无精症、严重寡精症患者的染色体异常和Y染色体微缺失

染色体异常和Y染色体微缺失被认为是两个白种人群中常见的生精障碍相关遗传因素。为了解中国无精症、严重寡精症患者中的染色体异常和Y染色体微缺失,运用染色体G显带技术,在358个原发无精症（256人）和严重寡精症（102人）不育患者中进行染色体核型分析;同时运用多重PCR技术,在核型正常的患者和100个正常生育男性中,对Y染色体AZF区微缺失进行筛查。在358个患者中,39人（10．9％）发现有染色体异常,Klinefelter（47,XYY）最为常见。无精症患者性染色体异常频率明显高于严重寡精症患者（12．1％VS1％）。在319个核型正常的患者中,46（14．4％）发现有AZF区微缺失,无精症和寡精症患者中Y染色体微缺失频率分别为15％和13．1％,AZFc区的微缺失最为常见,AZFa区的微缺失只见于无精症患者,正常生育男性中未发现AZF区的微缺失。结果显示,在中国无精症、严重寡精症患者中,大约25％的患者有染色体异常或Y染色体AZF区微缺失,提示这两种遗传异常是中国人群生精障碍的重要相关遗传病因,有必要在男性不育的诊断以及利用细胞浆内精子注射技术进行辅助生育时,对患者的这些遗传异常进行筛查。     

Purifying Selection Causes Widespread Distortions of Genealogical Structure on the Human X Chromosome

The extent to which selective forces shape patterns of genetic and genealogical variation is unknown in many species. Recent theoretical models have suggested that even relatively weak purifying selection may produce significant distortions in gene genealogies, but few studies have sought to quantify this effect in humans. Here, we employ a reconstruction method based on the ancestral recombination graph to infer genealogies across the length of the human X chromosome and to examine time to most recent common ancestor (TMRCA) and measures of tree imbalance at both broad and very fine scales. In agreement with theory, TMRCA is significantly reduced and genealogies are significantly more imbalanced in coding regions and introns when compared to intergenic regions, and these effects are increased in areas of greater evolutionary constraint. These distortions are present at multiple scales, and chromosomal regions as broad as 5 Mb show a significant negative correlation in TMRCA with exon density. We also show that areas of recent TMRCA are significantly associated with the disease-causing potential of site as measured by the MutationTaster prediction algorithm. Together, these findings suggest that purifying selection has significantly distorted human genealogical structure on both broad and fine scales and that few chromosomal regions escape selection-induced distortions.     

DXS6804/DXS9896/GATA144D04基因座在中国汉族群体中的遗传多态性及其法医学应用

为了调查X染色体上DXS6804、DXS9896和GATA144D04等3个STR基因座在中国汉族群体的遗传多态性及其法医学应用价值,来用PCR和聚丙烯酰胺凝胶电泳对X染色体3个STR基因座进行分型,并检验女性基因型频率分布是否符合Hardy Weinberg平衡,计算法医学常用各种概率。DXS6804、DXS9896和GATA144D04的非父排除率分别为0 5990、0 6220、0 4280,表明3个STR基因座在中国汉族群体均具有遗传多态性,χ2检验表明女性的基因型频率分布符合Hardy Weinberg平衡。X染色体上的基因座DXS6804、DXS9896和GATA144D04在中国汉族群体中具有较高的遗传多态性,可应用于法医学检验和群体遗传学分析。     

DXS6804/DXS9896/GATA144D04基因座在中国汉族群体中的遗传多态性及其法医学应用Genetic

为了调查X染色体上DXS6804、DXS9896和 GATA144D04等3个STR基因座在中国汉族群体的遗传多态性及其法医学应用价值,来用PCR和聚丙烯酰胺凝胶电泳对X染色体3个STR基因座进行分型,并检验女性基因型频率分布是否符合Hardy-Weinberg平衡,计算法医学常用各种概率。DXS6804、DXS9896和 GATA144D04的非父排除率分别为0.5990、0.6220、0.4280,表明3个STR基因座在中国汉族群体均具有遗传多态性,χ2检验表明女性的基因型频率分布符合Hardy-Weinberg平衡。X染色体上的基因座DXS6804、DXS9896和 GATA144D04在中国汉族群体中具有较高的遗传多态性,可应用于法医学检验和群体遗传学分析。 Abstract： To investigate the genetic polymorphisms of three short tandem repeats loci of chromosome X in Chinese Han population in Chengdu area and its use in forensic science. Three X-chromosome linked short tandom repeat loci were analyzed by PCR followed by polyacrylamide gel electrophoresis. Hardy-Weinberg equilibrium was tested and forensic interested value was calculated .The power of exlcution of DXS6804、DXS9896和 GATA144D04 is 0.5990、0.6220、0.4280,respectively. The result showed that all the three STR loci were polymorphic among 100 unrelated females and 120 unrelated males from Chinese Han population. χ2 tests demonstrated that genotype frequencies in females did not depart from Hardy-Weinberg equilibrium. Three X-chromosome linked short tandem repeat loci have high polymorphism, they can be applied to forensic medicine and population genetics.     

Evidence for Local Regulatory Control of Escape from Imprinted X Chromosome Inactivation

X chromosome inactivation (XCI) is an epigenetic process that almost completely inactivates one of two X chromosomes in somatic cells of mammalian females. A few genes are known to escape XCI and the mechanism for this escape remains unclear. Here, using mouse trophoblast stem (TS) cells, we address whether particular chromosomal interactions facilitate escape from imprinted XCI. We demonstrate that promoters of genes escaping XCI do not congregate to any particular region of the genome in TS cells. Further, the escape status of a gene was uncorrelated with the types of genomic features and gene activity located in contacted regions. Our results suggest that genes escaping imprinted XCI do so by using the same regulatory sequences as their expressed alleles on the active X chromosome. We suggest a model where regulatory control of escape from imprinted XCI is mediated by genomic elements located in close linear proximity to escaping genes.     

孤雌胚胎干细胞基因印迹及X染色体失活状态初步研究

目的:研究孤雌胚胎干细胞(phESC)与受精卵来源胚胎干细胞(hESC)在印迹基因表达、X染色体失活等方面的异同。方法:运用实时荧光相对定量PCR、甲基化特异性PCR和免疫荧光染色等方法检测phESC与hESC在父系印迹基因IGF2R,母系印迹基因SNRPN,IGF2相对表达量及X染色体失活状态。结果:①母系印迹基因SNRPN,IGF2在phESC细胞中不表达,而父系印迹基因IGF2R表达量则相对于hESC有近2倍的上调;②XIST基因在第35代phESC细胞中没有表达,意味着早期的phESC没有进行X染色体失活,而到了第55代,XIST基因开始表达并随着分化时间的延长表达量逐渐上调;③XIST启动子甲基化状态及组蛋白H3赖氨酸27三甲基化免疫荧光染色阳性证实phESC在长期培养后启动了X染色体失活。结论:phESC的X染色体失活状态在培养过程中存在不稳定的情况,建议对phESC进行更深入的表观遗传稳定性研究,以确保这种细胞未来安全、高效的应用。     

Sex- and Gamete-Specific Patterns of X Chromosome Segregation in a Trioecious Nematode

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