水稻配子体细胞质雄性不育基因orf79/orfH79的变异多态性研究

线粒体基因组异常嵌合基因orf79/orfH79被证实为水稻配子体细胞质雄性不育（cytoplasmic male sterility,CMS）基因。为了调查配子体CMS基因orf79/orfH79在水稻资源中的遗传与变异,来自不同国家的31份水稻材料被用于PCR检测。10份水稻材料被检测出具有配子体CMS基因orf79/orfH79,表明配子体CMS基因orf79/orfH79在水稻资源中具有较高的分布频率（32.2%）。DNA序列分析显示水稻配子体CMS基因orf79/orfH79具有非常保守的遗传特性（98.3%）,只有4个多态性碱基位点被检测出,4个碱基位点的变异导致多肽ORF79/ORFH79三个氨基酸位点的改变。基于orf79/orfH79 DNA序列的聚类分析系显示10份水稻材料被分成了3个类群,表明配子体CMS胞质在水稻资源中存在多种变异模式。研究结果为新水稻CMS胞质的发掘与培育提供重要的理论和实验依据。     

绵羊(0vis aries)线粒体DNA的遗传变异类型研究

通过PCR-RFLP和DNA序列测定发现绵羊线粒体基因组内存在的HinfⅠ酶切多态是COⅠ基因第234位的T-C碱基替换的结果.DNA序列分析表明该位点碱基的替换不导致氨基酸的改变,为一同义突变.依据这一单核苷酸多态可直接进行绵羊线粒体基因组的分型鉴定,并可作为绵羊核质基因互作研究、核外基因效应分析及转基因、动物克隆研究中胚胎及个体识别的分子标记.     

水稻的4个NADP-ME基因具有不同的表达模式

在高等植物中,NADP-苹果酸酶(NADP-ME)由一个小的基因家族编码.根据发表的水稻基因组序列信息,对水稻的NADP-ME家族进行了研究.结果表明:水稻NADP-ME家族由3个胞质型NADP-ME和1个质体型NADP-ME构成,发现两个新的胞质型NADP-ME基因. 尽管4个水稻NADP-ME的氨基酸序列存在较高的相似性,但是其中一个胞质型NADP-ME(OscytME3)在NADP-ME氨基酸序列保守区存在着不同于其他NADP-ME的特征. 进化树分析表明它可能起源于不同于其他NADP-ME的另一条进化分支.虽然4个NADP-ME基因表达的组织特异性和发育阶段特异性不尽相同,但是在功能上可能都参与植物的胁迫应答.     

石刁柏雄性连锁的AFLP及SCAR标记的建立

目的:对石刁柏雌雄株基因组差异进行分析,筛选雄性或雌性连锁的分子标记.方法:利用限制性片段长度多态性技术,设计了多个引物组合,分别对石刁柏雌雄株基因组进行扩增.结果:在使用的72个引物组合中,引物组合E-AAG+M-CAT从雄性基因组中扩增出了一个雄性连锁的标记(MLDA555),该序列长度为555bp,AT含量为59%.Blast检索未发现相似序列.根据该片段序列设计的引物将该标记转化为雄性连锁的大小为523bp的稳定的SCAR标记,经过不同基因型雄性个体的验证证明该标记广泛存在于不同基因型石刁柏雄性个体中.结论:通过AFLP扩增筛选得到了石刁柏雄性连锁的AFLP和SCAR标记,为石刁柏性别决定机制的理解及石刁柏的分子标记辅助育种提供理论资料和技术支持.     

石刁柏雄性连锁的AFLP及SCAR标记的建立

目的:对石刁柏雌雄株基因组差异进行分析,筛选雄性或雌性连锁的分子标记.方法:利用限制性片段长度多态性技术,设计了多个引物组合,分别对石刁柏雌雄株基因组进行扩增.结果:在使用的72个引物组合中,引物组合E-AAG+M-CAT从雄性基因组中扩增出了一个雄性连锁的标记(MLDA555),该序列长度为555bp,AT含量为59%.Blast检索未发现相似序列.根据该片段序列设计的引物将该标记转化为雄性连锁的大小为523bp的稳定的SCAR标记,经过不同基因型雄性个体的验证证明该标记广泛存在于不同基因型石刁柏雄性个体中.结论:通过AFLP扩增筛选得到了石刁柏雄性连锁的AFLP和SCAR标记,为石刁柏性别决定机制的理解及石刁柏的分子标记辅助育种提供理论资料和技术支持.     

基于线粒体基因组全序列的鳅科(Cobitidae)鱼类进化关系分析

为了解鳅科(Cobitidae)鱼类线粒体全基因组序列的结构特征和系统发育信息,利用生物信息学方法对已知的40种鳅科鱼类线粒体全基因组进行比对分析,用邻接法(NJ)构建系统发育树。结果显示：(1)鳅科鱼类线粒体基因组全序列长度在16 553～16 937 bp之间,基因组的结构特征及基因排列顺序与其他硬骨鱼类一致。(2)鳅科鱼类线粒体全基因组一致序列长度为17 483 bp,变异位点数为8039(45.9%),Kimura双参数平均遗传距离为0.19。在13个蛋白质编码基因中,ND2的变异程度(58.9%)和平均遗传距离(0.28)最大,变异程度最小的是12S rRNA(30.5%),tRNA拼接序列的平均遗传距离最小(0.07)。(3)Cox1、ND5、Cytb基因是进行鳅科鱼类系统发育分析较为理想的分子标记,NJ系统发育树显示,条鳅亚科(Noemacheilinae)是最早分化出来的一枝群系,位于祖先位置,沙鳅亚科(Botiinae)和花鳅亚科(Cobitinae)亲缘关系更近,互为姐妹群系。本研究为鳅科鱼类进化学研究和分子标记的选取提供参考依据。     

DNA分子标记在不结球白菜遗传育种中的应用进展

概述了分子标记的分类和特点,以及在不结球白菜遗传多样性分析、遗传图谱构建、数量性状座位(quantitative trait locus,QTLs)定位和分子标记辅助育种等领域中的最新研究进展.总结了目前分子标记在不结球白菜遗传改良和辅助育种中存在的若干问题,并对该研究领域提出了一些展望和建议.     

苏姜猪线粒体基因组全长测序及线粒体基因多样性分析

从分子水平上探究苏姜猪的遗传多样性,采用二代测序方法测定了苏姜猪线粒体基因组全序列,整合NCBI数据库中已公布的家猪类,以及本实验室测定的苏姜猪线粒体基因组全序列,使用蛋白编码基因和rRNA基因联合的数据集,基于贝叶斯法(BI法)和最大似然法(ML法)构建了家猪类的系统发生树,测序结果表明,该样本线粒体基因组全长15 436 bps,包含典型的家猪类线粒体基因组37条基因,结构紧凑,其线粒体基因组碱基组成(A+T)约占40%,(C+G)约占60%,t RNA总共有8处错配,结果表明苏姜猪具有丰富的遗传多样性,江海型线粒体核苷酸多态性高于全国6个类型猪的平均水平,体现了其独特的遗传多样性.在基于线粒体基因组完整编码区的系统发育分析中,苏姜猪在中国6个类型家猪中单独成为一支,支持其作为一个新猪种而存在.     

绵羊（Ovis aries）线粒体DNA的遗传变异类型研究

通过PCR－RFLP和DNA序列测定发现绵羊线粒体基因组内存在的HinfⅠ酶切多态是COⅠ基因第234位的T－C碱基替换的结果。DNA序列分析表明该位点碱基的替换不导酸的改变，为一同义突变。依据这一单核苷酸多态可直接进行绵羊线粒体基因组的分型鉴定，并可作为绵羊核质基因互作研究、核外基因效应分析及转基因、动物克隆研究中胚胎及个体识别的分子标记。     

波纹鳜线粒体基因克隆及系统发育信息分析

为了探讨鳜类线粒体基因组结构特征及系统发育信息,以期为鳜类遗传学研究和分子标记提供参考依据,本文将波纹鳜Siniperca undulata线粒体随机切割成长度为1～3 kb的15个片段,采用PCR产物直接测序法获得波纹鳜线粒体全基因组,并提交至NCBI数据库,获得登录号KJ644783。结果分析发现:波纹鳜线粒体包含13个编码蛋白基因、22个tRNA、2个rRNA以及一个超变区(D-loop),基因排列顺序与其他硬骨鱼类似,不存在基因重排现象。整个基因组存在明显的AC偏好性,其中G碱基含量非常低,仅仅为16.23%。系统发育信息分析结果显示,CO1、ND5、CYTB、ND4基因具有较好的进化适应性,能够较好地反映鳜类的系统发育关系。进一步的进化分析显示,鳜类分成了鳜属和少鳞鳜属。     

不结球白菜种质创新及资源信息管理研究进展

对不结球白菜(Brassica campestris ssp. chinensis Makino)抗性育种、细胞质雄性不育、基因克隆和转基因育种、遗传图谱和DNA分子标记辅助育种,以及种质资源信息管理进行了综述,以期为不结球白菜种质创新和管理提供参考和建议.     

Construction of a bacterial artificial chromosome library of S-type CMS maize mitochondria

In order to isolate mitochondrial genes easily, we have developed a new method to construct S-type CMS maize mitochondrial gene library by means of embedding mitochondria and enzymatic digesting mitochondriain situ, preparing mtDNA by electrophoresis, digesting LMP agarose with β-agarase, using BAC vector and electroporation. About 2 500 white clones of Mo17 CMS-J mitochondrial gene library were obtained with the average size of 18.24 kb, ranging from 5 to 40 kb, 63.6% inserts came from mitochondrial genome and represented 48 × mitochondrial genome equivalents. All the probes had detected the positive clones in the gene library. It is helpful to elucidating the maize mitochondrial genome structure and mechanism of S-type CMS, and may give some valuable reference to the construction of other plant mitochondrial genome library.     

红莲型水稻不育系与保持系线粒体DNA酶切分析

为研究红莲型水稻细胞质雄性不育的分子基础，我们提取纯化了红莲型水稻丛广４１Ａ和丛广４１Ｂ的线粒体ＤＮＡ，采用限制性内切酶酶切片段长度多态性（ＲＦＬＰ）方法比较了红莲型不育系和保持系线粒体ＤＮＡ酶切图谱，发现两者之间存在一定的差异，部分支持了细胞质雄性不育与线粒体ＤＮＡ有关的结论。     

Genetic typing of mitochondrial DNA in sheep (Ovis aries )

Using PCR-RFLP and DNA sequencing, this study confirmed that HinfI polymorphism in the ovine mitochondrial COI gene resulted from the T-C substitution at the nucleotide 234 but this mutation did not encode another amino acid, which was actually a synonymous mutation. This single nucleotide polymorphism can be used as gene typing marker for mitochondrial genome in the research of the interactions between mitochondria and nucleus, extranuclear gene effects, and as molecular discriminating marker for embryo or individual in the research area of gene transfer and animal cloning.     

林木分子育种研究的基因组学信息资源述评

分子育种是指利用与性状相关的DNA标记进行选育,也称标记辅助选择或标记辅助育种,广义上还包括基因工程育种和基因组学辅助育种。林木分子育种为早期选择和加速育种提供了极具潜力的高效手段。笔者对林木分子育种研究的基因组学信息资源进行了进展综述和前景展望。近30年来,林木分子标记技术从早期的低通量方法发展到目前基于微阵列芯片和新一代测序的高通量技术,如测序分型、转录组测序、重测序、扩增子测序和外显子组测序等,并广泛用于连锁作图、关联分析和基因组选择等林木性状相关的DNA变异检测研究。随着2006年毛果杨基因组序列的发表,已有50余个树种完成了基因组测序。基于连锁作图和关联研究检测了林木10余个属生长、材性和抗逆及非木质产品品质等性状相关的大量基因组位点,主要趋势表现为：① 表型广泛,涵盖经济性状、生理指标和代谢成分等;②标记数量成千上万甚至上百万,覆盖全基因组;③转录组和降解组等多组学的分子变异开始应用;④ 利用大群体以提高位点检测的精度;⑤ 重视环境的影响,大田试验设置多个地点,解析QTL与环境、年份的互作效应;⑥ 结合参考基因组序列和/或转录组差异表达基因进一步挖掘性状相关的候选基因,建立了桉属、松属和云杉属等主要造林树种的基因组选择模型。此外,积累了泛基因组、相关软件和算法、功能基因、基因组编辑技术及网站和数据库等其他信息资源。林木分子育种面临的挑战主要包括：① 如何获得稳定性好的性状相关基因组位点和基因组选择（GS）模型;② 缺乏自动化、无损和高通量的表型测定技术;③对大基因组的针叶树和一些多倍体树种,仍难获得高质量的基因组序列;④ 标记辅助选择增加了常规育种之外的费用,且存在不确定性;⑤多数树种的加速育种仍较困难。后基因组时代的林木分子育种将有效结合到常规育种程序中,显著促进遗传增益的提高。     

The genetic and molecular basis of cytoplasmic male sterility and fertility restoration in rice

Cytoplasmic male sterility （CMS） is a maternally inherited characteristic found in many （〉150） plant species. CMS/restoration systems are useful tools for hybrid seed production, and are ideal models for study of the interactions between nuclear and mitochondrial genomes. CMS/restoration systems in rice have been widely used for hybrid seed production, greatly contributing to the food supply. This article reviews the progress of the studies on the genetic and molecular basis of cytoplasmic male sterility and fertility restoration in rice.     

Sequence analysis of the gene correlated with cytoplasmic male sterility (CMS) in rape-seed (Brassica napus) Polima and Shaan 2A

orf224 is a CMS-related mitochondrial gene discovered in Polima cytoplasm. Shaan 2A CMS line is the parent of the first rapeseed hybrid cultivar Qinyou No. 2that has been grown in many regions of China. In this work,genomic DNA of Polima CMS line and Shaan 2A CMS line were used as templates, two primers of specific oligonucleoamplification fragments were cloned into pGEM-T Easy vectors and DNA sequences were determined. The CMSassociated gene, orf224-1 present in Shaan 2A CMS line, has a sequence highly homologous to the orf224 of the Polima CMS line, except for one nucleotide at position +398. There were only one base (AAC→AGC) and one amino acid (Asn →Ser) differences between the two. The homologies of the two sequences in nucleotide and. amino acid were 99.9 % and 99.6%, respectively. It is concluded that orf224 in Polima CMS line and orf224-1 of Shaan 2A CMS line are the allele at the same locus in mitochondria.``     

A DNA sequence alignment algorithm using quality information and a fuzzy inference method

DNA sequence alignment algorithms in computational molecular biology have been improved by diverse methods. In this paper, we propose a DNA sequence alignment that uses quality information and a fuzzy inference method developed based on the characteristics of DNA fragments and a fuzzy logic system in order to improve conventional DNA sequence alignment methods that uses DNA sequence quality information. In conventional algorithms, DNA sequence alignment scores are calculated by the global sequence alignment algo- rithm proposed by Needleman-Wunsch, which is established by using quality information of each DNA fragment. However, there may be errors in the process of calculating DNA sequence alignment scores when the quality of DNA fragment tips is low, because only the overall DNA sequence quality information are used. In our proposed method, an exact DNA sequence alignment can be achieved in spite of the low quality of DNA fragment tips by improvement of conventional algorithms using quality information. Mapping score param- eters used to calculate DNA sequence alignment scores are dynamically adjusted by the fuzzy logic system utilizing lengths of DNA fragments and frequencies of low quality DNA bases in the fragments. From the experiments by applying real genome data of National Center for Biotechnology Information, we could see that the proposed method is more efficient than conventional algorithms.     

A DNA sequence alignment algorithm using quality information and a fuzzy inference method

DNA sequence alignment algorithms in computational molecular biology have been improved by diverse methods. In this paper, we propose a DNA sequence alignment that uses quality information and a fuzzy inference method developed based on characteristics of DNA fragments and a fuzzy logic system in order to improve conventional DNA sequence alignment methods that uses DNA sequence quality information. In conventional algorithms, DNA sequence alignment scores are calculated by the global sequence alignment algorithm proposed by Needleman-Wunsch, which is established by using quality information of each DNA fragment. However, there may be errors in the process of calculating DNA sequence alignment scores when the quality of DNA fragment tips is low, because only overall DNA sequence quality information are used. In our proposed method, an exact DNA sequence alignment can be achieved in spite of low quality of DNA fragment tips by improvement of conventional algorithms using quality information. Mapping score parameters used to calculate DNA sequence alignment scores are dynamically adjusted by the fuzzy logic system utilizing lengths of DNA fragments and frequencies of low quality DNA bases in the fragments. From the experiments by applying real genome data of National Center for Biotechnology Information, we could see that the proposed method is more efficient than conventional algorithms.