Dissecting Arabidopsis thaliana DICER function in small RNA processing, gene silencing and DNA methylation patterning.

Small RNAs have several important biological functions. MicroRNAs (miRNAs) and trans-acting small interfering RNAs (tasiRNAs) regulate mRNA stability and translation, and siRNAs cause post-transcriptional gene silencing of transposons, viruses and transgenes and are important in both the establishment and maintenance of cytosine DNA methylation. Here, we study the role of the four Arabidopsis thaliana DICER-LIKE genes (DCL1-DCL4) in these processes. Sequencing of small RNAs from a dcl2 dcl3 dcl4 triple mutant showed markedly reduced tasiRNA and siRNA production and indicated that DCL1, in addition to its role as the major enzyme for processing miRNAs, has a previously unknown role in the production of small RNAs from endogenous inverted repeats. DCL2, DCL3 and DCL4 showed functional redundancy in siRNA and tasiRNA production and in the establishment and maintenance of DNA methylation. Our studies also suggest that asymmetric DNA methylation can be maintained by pathways that do not require siRNAs.

Genome-wide high-resolution mapping and functional analysis of DNA methylation in arabidopsis.

Cytosine methylation is important for transposon silencing and epigenetic regulation of endogenous genes, although the extent to which this DNA modification functions to regulate the genome is still unknown. Here we report the first comprehensive DNA methylation map of an entire genome, at 35 base pair resolution, using the flowering plant Arabidopsis thaliana as a model. We find that pericentromeric heterochromatin, repetitive sequences, and regions producing small interfering RNAs are heavily methylated. Unexpectedly, over one-third of expressed genes contain methylation within transcribed regions, whereas only approximately 5% of genes show methylation within promoter regions. Interestingly, genes methylated in transcribed regions are highly expressed and constitutively active, whereas promoter-methylated genes show a greater degree of tissue-specific expression. Whole-genome tiling-array transcriptional profiling of DNA methyltransferase null mutants identified hundreds of genes and intergenic noncoding RNAs with altered expression levels, many of which may be epigenetically controlled by DNA methylation.

鮡科褶鮡属鱼类部分线粒体DNA序列分析与分子进化

采用PCR技术获得了中国鮡科褶鮡属(Pseudecheneis)5种鱼类及外群种类巨魾[Bagarius yarrelli(Sykes)]和红河纹胸鮡(Glyptothorax fukiensis honghensis Li)的线粒体部分基因序列.序列分析结果表明:间褶鮡(Pseudecheneis intermedius Chu)与平吻褶鮡(P.paviei Vaillant)在Cyt b基因片段上完全无差异,形成单倍型,支持间褶鮡应为平吻褶鮡的同物异名.凭Cyt b基因片段构建了它们的NJ,MP和ML分子树,3棵分子树基本一致,均支持褶鮡属构成一单系群;怒江和伊洛瓦底江"黄斑褶鮡"不同样品分别聚在一起,但二水系的样品未能形成一单系;其他各种的不同样品均能分别聚在一起形成单系.怒江和伊洛瓦底江"黄斑褶鮡"的分类地位值得今后进一步研究.

Tandem repeats upstream of the Arabidopsis endogene SDC recruit non-CG DNA methylation and initiate siRNA spreading.

Plants use siRNAs to target cytosine DNA methylation to both symmetrical CG and nonsymmetrical (CHG and CHH) sequence contexts. DNA methylation and siRNA clusters most frequently overlap with transposons in the Arabidopsis thaliana genome. However, a significant number of protein-coding genes also show promoter DNA methylation, and this can be used to silence their expression. Loss of the majority of non-CG DNA methylation in drm1 drm2 cmt3 triple mutants leads to developmental phenotypes. We identified the gene responsible for these phenotypes as SUPPRESSOR OF drm1 drm2 cmt3 (SDC), which encodes an F-box protein and possesses seven promoter tandem repeats. The SDC repeats show a unique silencing requirement for non-CG DNA methylation directed redundantly by histone methylation and siRNAs, and display spreading of siRNAs and methylation beyond the repeated region. In addition to revealing the complexity of DNA methylation control in A. thaliana, SDC has important implications for how plant genomes utilize gene silencing to repress endogenous genes.

RNAi, DRD1, and histone methylation actively target developmentally important Non-CG DNA methylation in Arabidopsis

Cytosine DNA methylation protects eukaryotic genomes by silencing transposons and harmful DNAs, but also regulates gene expression during normal development. Loss of CG methylation in the Arabidopsis thaliana met1 and ddm1 mutants causes varied and stochastic developmental defects that are often inherited independently of the original met1 or ddm1 mutation. Loss of non-CG methylation in plants with combined mutations in the DRM and CMT3 genes also causes a suite of developmental defects. We show here that the pleiotropic developmental defects of drm1 drm2 cmt3 triple mutant plants are fully recessive, and unlike phenotypes caused by met1 and ddm1, are not inherited independently of the drm and cmt3 mutations. Developmental phenotypes are also reversed when drm1 drm2 cmt3 plants are transformed with DRM2 or CMT3, implying that non-CG DNA methylation is efficiently re-established by sequence-specific signals. We provide evidence that these signals include RNA silencing though the 24-nucleotide short interfering RNA (siRNA) pathway as well as histone H3K9 methylation, both of which converge on the putative chromatin-remodeling protein DRD1. These signals act in at least three partially intersecting pathways that control the locus-specific patterning of non-CG methylation by the DRM2 and CMT3 methyltransferases. Our results suggest that non-CG DNA methylation that is inherited via a network of persistent targeting signals has been co-opted to regulate developmentally important genes. © 2006 Chan et al.

Mitochondrial DNA D2loop Variation and Genetic Background of Brahman Cattle

对10 头原种婆罗门牛mtDNA D2loop 全序列912 bp 测序, 婆罗门牛遗传多样性丰富, 检测到的9 种单 倍型兼有瘤牛( B . indicus) 与普通牛( B . taurus) 的遗传背景, 核苷酸变异率为6125 % , 单倍型多态度为01978 ± 01054 , 核苷酸多态度为01014 30 ±01008 68。所有单倍型聚为明显的两大分支, 婆罗门牛的大部分单倍型为普通 牛单倍型类群, 并占绝对优势(90 %) , 仅Brah26 与亚洲瘤牛聚在一起, 属于亚洲瘤牛线粒体单倍型, 表明婆罗门 牛的确是集亚洲瘤牛、欧洲普通牛等优良特性于一身(易产犊、产肉性能好、耐热与体表寄生虫等) 的瘤牛品种之 一。育种学家引种瘤牛的目的是改善当地牛的生产力与适应性, 现代普通牛表现出明显又普遍的瘤牛渐渗现象。 对现代的瘤牛品种而言, 除亚洲瘤牛品种外, 普通牛对其他瘤牛品种育成的贡献同样高。支持瘤牛( B . indicus ) 为独立驯化、起源于印度次大陆的假说。

线粒体DNA 3243、3316 点突变与2 型糖尿病

　目的　研究2 型糖尿病患者中线粒体tRNAL eu (UUR) 基因3243AöG 突变和NADH 脱氢酶亚 单位1 基因( ND1 ) 基因3316GöA 突变的发生频率及其与2 型糖尿病的相关性。方法　应用聚合酶链反 应及限制性片段长度多态性技术检测225 例中国云南2 型糖尿病患者和195 名无糖尿病家族史的健康对 照者有无3243AöG 突变和3316GöA 突变, 并经DNA 直接测序确证。结果　2 型糖尿病患者中3316GöA 突变者5 例(2. 22% ) , 195 例对照者中突变者2 例(1. 03% ) , 突变发生率在两组间差异无统计学意义(P = 0. 4576) ; 两组中无线粒体3243AöG 突变。结论　线粒体tRNAL eu (UUR) 基因3243AöG 突变在中国云南2 型 糖尿病人群中发生频率低, 可能不是云南人群中2 型糖尿病的常见病因。线粒体ND1 基因3316GöA 突变 可能仅为人群中线粒体基因组的正常多态。其他的遗传、环境及子宫内因素需要进一步研究。

Molecular Phylogeny of Slow Lorises ( Nycticebus) Revealed by D2loop Sequences and Complete Cytochrome bGene Sequences of Mitochondrial DNA

本研究测定了懒猴属( Nycticebus) D 环的部分序列和细胞色素b 基因的全序列(1 140 bp) , 分析了 该属物种之间的系统发育进化关系。在DNA 水平上, 序列分析结果一致地提供了新的分类学证据: 支持Rata2 jszczak 和Groves 的观点, 即N1intermedus 只是N1 pygmaeus 的成体(Ratajszczak , 1998 ; Groves , 1971) 。对两种 序列的数据做了联合及个别分析, 获得相似的系统树, 支持懒猴属由两个单系群组成: 第一群由N1 pygmaeus 聚成, 第二群由N1coucang 聚成。该结果也提供了新的分子遗传证据, 支持懒猴属由N1coucang 和 N1 pygmaeus 两物种组成。

Study on Mitochondrial DNA Cytochrome b Gene of Chinese sea bass, Lateolabrax sp.

参考鳗鲡等鱼类线粒体 DNA序列进行了中国花鲈线粒体 DNA细胞色素 b基因片断的引物设计、PCR扩增及其序列测定。得到中国花鲈的碱基序列为 4 10 bp,其 A、T、G、C含量分别为 10 1bp(2 4 .6 3% )、112 bp(2 7.32 % )、72 bp(17.56 % )、12 5bp(30 .4 9% ) ,与鳗鲡等其他鱼类相同基因片断序列碱基含量相似。

7种长蠹科昆虫的线粒体DNA ND4基因序列比较分析

长蠹科昆虫严重危害林木和仓贮物品。该文应用非损伤性DNA测序技术测定了来自不同国家的长蠹科害虫的线粒体DNAND4基因的部分序列。在获得的 2 0 4bp的序列中 ,7种昆虫的序列变异丰富 ,多数变异发生在密码子的第 3位点上。将实验结果与形态学特征比较分析 ,探讨 7个种的形态差异与基因序列的差异 ;结果表明 ,7种昆虫不仅在外形特征上存在差异 ,而且在ND4基因序列上的差异程度也很显著 ,平均为 2 1 94%。这为分类鉴定提供了充分的证据。

云南东方蜜蜂随机扩增多态DNA研究

本文利用随机扩增多枣DN^技术研究r云南 东方童蜂t工蜂)32个个体的遗传变异。帆25个叫物中缔 选出11十引物，其中9个引物扩增出事惫谱带。共检测到 63杀扩增片段，其中46条出现变异。用Nei的片断共事度 盐式计算了，2个个体的遗传距离，用㈣M^聚盎芒构营， 系统发育蝌状图。系统寨娄图中大多数采自同～地匹的样奉 聚在一起．但也存在一宅的交叉，提示地理群体H近期可能 存在一定程度的善因流。

中国民族人群线粒体DNA 9bp序列 缺失的分布

采用PCR—PAGE方法，对16个少数民族群体和9个地区的汉族群体共1521 人进行了线粒体基因组COIl和tRNAb8基因间非编码序列中串联重复的9 bp序列缺失情况的检测．贵州苗族、布依旅的缺失频率在所研究的民族人群中最高，分别达到 32·4％和30．8％，云南佤族、拉祜族和新疆维吾尔族、蒙古族频率相当低(≤4％)，新疆哈萨克族群体中未检测到缺失类型，而其他群体的频率表现为中等或较低(6％。 24％)·除新疆汉族、贵州汉族和报道的台湾汉族外，其他各地汉族群体间缺失频率差异不大；有些来自同一地区或共同历史起源的民族群体的缺失频率没有表现出相边的分布趋势·总体上南方的、沿海的民族群体比北方的、内地的群体表现出较高的缺失频率．综合统计已报道的和此次研究的中国人群的9 bp缺失频率约为17．20％．另外，还在4个个体中检测到并经测序证实存在9 bp的3个重复；在随机抽样选择测定的有缺失和没有妖失的27个个体中，仅在1例中发现有报道的X．II型9 bp模式，

云南鹅线粒体DNA多态及酶切图谱分析

用１８种限制性内切酶对７只云南鹅的线粒体ＤＮＡ（ｍｔＤＮＡ）进行了限制性片段长度多态（ＲＦＬＰ）分析。１７种酶在所有受试个体表现为一致的限制性态型（ｍｏｒｐｈ），一只灰羽鹅用ＰｖｕＩ消化时出现变异。用１４种限制性内切酶构建了云南鹅ｍｔＤＮＡ的酶切图谱。

贵州绵羊线粒体DNA (m tDNA ) 多态性研究

采用15 种限制性内切酶对贵州威宁绵羊和六盘水绵羊各5 只个体的m tD2 NA 多态性进行研究。其中只有BamHÉ 一种酶的酶切类型表现多态, 产生三种酶切 类型。共得到17 种限制性态型, 可归结为三种m tDNA 单倍型, 在威宁绵羊中单倍 型É 所占比较较大(80% ) , 单倍型Ë 只占20% , 而在六盘水绵羊中则单倍型É 和Ê 所占比例相当; 根据单倍型在群体中的分布和单倍型间遗传距离, 推测贵州绵羊的 m tDNA 单倍型É 和Ê 可能代表两个母系祖先的基本单倍型, 其分化时间约有16～ 32 万年之久, 单倍型Ë 则可能是由单倍型É 随机突变而来; 两个地方绵羊群体间的 分化时间大约为613～ 1216 万年, 群体间的m tDNA 遗传多态度较低, 但六盘水绵羊 群体内遗传多态度比威宁绵羊较丰富。

黔东南小香羊与贵州原有其它山羊品种的线粒体DNA 多态性比较

用B am H É、B g lÉ、B g lÊ、D raÉ、E coR É、E coR Í、H indË、Kp nÉ、 P stÉ、P vuÊ、S acÉ、S a lÉ、Sm aÉ、S tuÉ、X hoÉ 15 种识别6 碱基的限制性内 切酶对黔东南小香羊和贵州原有3 个地方山羊品种的93 只个体的m tDNA 进行分 析表明, B am H É、H indË 和S a lÉ 3 种酶表现多态性; 共检出18 种限制性多态型, 归纳得到3 种单倍型, 以单倍型É 和Ê 为基本单倍型。根据此2 种基本单倍型在所 比较各品种中的不同分布比例, 以及遗传距离分析和品种间的聚类关系, 表明黔东 南小香羊的群体遗传构成与贵州省原有其它3 个山羊品种不同, 从而为进一步确认 其为一独立的品种提供了必要的分子生物学依据。