基于线粒体Cytb基因序列的褐石斑鱼种群遗传多样性分析

为评估种质资源状况被世界自然保护联盟(International Union for Conservation of Nature, IUCN)评定为易危物种的褐石斑鱼(Epinephelus bruneus)野生种群的遗传多样性和遗传分化水平，采用PCR方法测定褐石斑鱼西太平洋海区的中国海南岛(HN)、福建厦门(XM)和韩国济州岛(HG)3个地理群体的线粒体细胞色素b(Cytb)基因的部分序列，并对其基因序列遗传变异、谱系结构和群体扩张历史特征进行分析。结果显示，褐石斑鱼3个地理种群(88个个体)共检测出18个多态位点，共有7种单倍型；各地理群体遗传多样性水平较低，而且单倍型在群体间分布不均，韩国群体遗传多样性最高，中国海南和厦门群体遗传多样性较低。地理距离最远的韩国群体和中国海南群体遗传分化最高(F_ST=0.177 5),地理距离最近的中国海南群体和厦门群体的遗传分化最低(F_ST=0.013 4)。Mantel检验结果显示，3个褐石斑鱼群体间遗传距离和地理距离间存在显著相关，距离隔离(Isolation by Distance, IBD)...     

基于ISSR数据探讨节茎曲柄藓(Campylopus umbellatus(Arn.) Paris)遗传多样性和居群遗传结构

选用简单序列重复区间扩增多态性(ISSR)分子标记,对分布于浙江清凉峰国家级自然保护区顺溪坞和凉源、浙江台州临海天台山、浙江舟山嵊泗列岛、江苏苏州天平山、广西上思十万大山、台湾宜兰县马告生态公园和云南西双版纳纳板河流域国家自然保护区8个自然居群的40份节茎曲柄藓(Campylopus umbellatus(Arn.) Par.)的遗传多样性进行了研究.应用筛选出的12条引物,共扩增出172条清晰、重复性高的条带.使用POPGENE 3.2分析发现:其中158条条带的多态性位点百分率(PPB)为91.86%,根井正利基因多样性指数(H)为0.292 2,香农-维纳多样性指数(I)为0.445 9,说明节茎曲柄藓居群遗传多样性水平较高.8个居群的遗传分化系数(G_(st))为0.634 6,居群间的基因流(N_m)为0.287 9,63.33%的遗传变异存在于居群间,36.67%的遗传变异存在于居群内,说明节茎曲柄藓居群间的遗传分化明显.基于ISSR数据的聚类分析表明:以遗传距离65为分组阈值,8个居群可被分为6组,节茎曲柄藓居群间的遗传分化主要由地理因素造成,居群内的遗传分化可能与生境的异质性有关.     

珍稀濒危植物石山苏铁的SSR引物设计和遗传多样性分析

揭示珍稀濒危植物石山苏铁(Cycas sexseminifera)种群的遗传多样性水平和遗传分化大小,确定优先保护种群,对石山苏铁的有效保护和管理策略的制定具有重要意义。本研究基于简化基因组序列分析设计Simple Sequence Repeats (SSR)引物,并利用SSR引物进行遗传多样性检测。实验共获得6对SSR引物,6对引物共检测出37个观测等位基因(Na),其中最小观测等位基因数目为3,最大观测等位基因数目为12,平均每个位点等位基因数目为6.167。位点GZST002、GZST055、GZST065、GZST088是高度多态性位点(PIC>0.5)。石山苏铁各种群期望杂合度(He)为0.330-0.533,平均值为0.444,表明石山苏铁遗传多样性处于中等水平;7个种群的He值大小顺序为广西植物研究所引种(ZWS)>崇左广河(GH)>崇左排汝(PR)>隆安陇怀(LH)>崇左中干(ZG)>百色作登(ZD)>隆安新会(XH)。固定系数(F)为-0.161-0.480,平均值为0.074,其中LH、PR、XH 3个野生种群为负值,GH、ZD、ZG 3个野生种群的固定系数则大于0;7个种群的F值大小顺序为百色作登(ZD)>广西植物研究所引种(ZWS)>崇左中干(ZG)>崇左广河(GH)>隆安陇怀(LH)>隆安新会(XH)>崇左排汝(PR)。因此,综合各个遗传指标,石山苏铁需要重点保护和引种的野生种群是遗传多样性相对较高的崇左广河(GH)种群。同时建议开发更多的SSR引物和采集更广范围的石山苏铁野生种群,精准筛选遗传多样性高的优良种群,以利于更全面和准确地评估石山苏铁的遗传多样性水平和濒危机制。     

基于SSR分子标记的广西极小种群野生植物长叶苏铁遗传多样性分析

了解广西区内的国家一级重点保护植物长叶苏铁（Cycas dolichophylla K.D.Hill）的遗传多样性和变异特征对广西长叶苏铁的保护和管理有着重要意义。本研究利用筛选出的6对稳定性好的SSR引物对分别来自广西百色市田阳县梅茶镇（PL）、德保县那甲镇（NL）和德保县城关镇（QH）的3个长叶苏铁种群的遗传多样性和变异特征进行分析。结果表明：6对引物中,高度多态性引物（GZST055、GZST065、GZST088）和低度多态性引物（GZST002、GZST013、GZST019）各占50%。广西长叶苏铁3个种群的平均观测等位基因为2.833,平均有效等位基因为2.005,平均Shannon信息指数为0.657,平均观测杂合度为0.259,平均期望杂合度为0.359。群体的遗传分化系数为0.049,说明群体间遗传分化小,且群体内遗传变异占所有遗传变异的97%,3个种群的基因交流频繁且遗传背景相互混杂。总体来看,广西长叶苏铁遗传多样性水平较低,而德保县那甲镇（NL）种群的遗传多样性相对最高,应将该种群作为重点保护单元进行保护。本研究还对广西长叶苏铁资源保护工作提出了建议和对策。     

Fine mapping of the red plant gene R1 in upland cotton(Gossypium hirsutum)

Sub 16 is a substitution line with G. hirsutum cv. TM-1 genetic background except that the 16th chromosome (Chr. 16) is replaced by the corresponding homozygous chromosome of G. barbadense cv. 3-79, and T586 is a G. hirsutum multiple gene marker line with 8 dominant mutation genes. The R ₁ gene for anthocyanin pigmentation was tagged in Chr. 16 in T586. The objective of this research was to screen SSR markers tightly linked with R ₁ by using the F₂ segregating population containing 1259 plants derived from the cross of Sub 16 and T586 and the backbone genetic linkage map from G. hirsutum×G. barbadense BC₁ newly updated by our laboratory. Genetic analysis suggested that the segregation ratio of red plants in the F₂ population fit Mendelian 1:2:1 inheritance, confirming that the red plant trait was controlled by an incomplete dominance gene. Preliminary mapping of R ₁ was conducted using 237 randomLy selected F₂ individuals and JoinMap v3.0 software. Then, a fine map of R1 was constructed using the F₂ segregating population containing 1259 plants, and R ₁ was located between NAU4956 and NAU6752, with only 0.49 cM to the nearest maker loci (NAU6752). These results provided a foundation for map-based cloning of R ₁ and further development of cotton cultivars with red fibers by transgenic technology. Supported by National Natural Science Foundation of China (Grant No. 30730067) and Programme of Introducing Talents of Discipline to Universities (Grant No. B08025)     

Rapid establishment of pure lines of silver carp (Hypophthalmichthys molirix) and bighead carp (Aristichthys nobilis)

The diversity of gynogenetic, artificial sex reversal and natural silver carp and bighead carp is examined using randomly amplified polymorphic DNA (RAPD) method.All of the 187 bands are obtained and 19 (10.16%) of them are polymorphic in gynogenetic silver carp.Meanwhile 32 (15.61%) out of 205 bands are polymorphic in control group.In gynogenetic bighead carp a total of 232 bands are identified and 11 (4.74%) out of them are polymorphic, while 25 (10.37%) out of 241 bands are polymorphic in control group.The genetic distance of four populations is calculated and it is 0.102 and 0.023 for gynogenetic silver carp and gynogenetic bighead carp respectively.The values of natural silver carp and bighead carp are 0.161 and 0.104.From the UPGMA trees constructed based on genetic distance, the sex reversal individuals that match with the gynogenetic female individuals are picked out.A new breeding process of establishing a pure line is developed.     

基于SSR的广西同色兜兰和带叶兜兰遗传多样性分析

评估四季开花型同色兜兰（Paphiopedilum concolor）和春季开花型带叶兜兰（P.hirsutissimum）的遗传多样性和种群结构,探究不同开花习性兜兰属物种间的遗传分化,为兜兰的可持续利用和野生资源保护提供理论依据。本研究利用10个多态性EST-SSR位点对12个同色兜兰和带叶兜兰种群的231个个体分型,开展遗传多样性、瓶颈效应、系统发育、遗传结构、遗传分化、距离隔离和环境隔离分析。结果表明,同色兜兰种群的大多数遗传多样性指标都显著（P<0.05）低于带叶兜兰;分子方差分析（AMOVA）结果显示,两者来自种群间的变异都较少,且带叶兜兰来自个体内的遗传变异高于同色兜兰。在同色兜兰的两个种群中检测到显著（P<0.05）瓶颈效应,而在带叶兜兰全部种群中均未检测到。群体水平的非加权平均（UPGMA）树和个体水平的邻接（NJ）树均显示同色兜兰和带叶兜兰间系统发育关系相对较近,并各自聚为一支。主坐标分析（PCoA）和Structure分析结果均支持系统发育分析结果。此外,遗传分化系数（F_ST）证实两个物种间的种群分化已达到较高水平。狭窄的自然分布和过去的种群瓶颈可能导致了同色兜兰的部分遗传多样性丧失。地理隔离是驱动同色兜兰和带叶兜兰种群遗传分化的主要因素,而环境隔离的作用较小。     

蝗总科6种蝗虫6个种群的遗传分化

采用水平淀粉凝胶电泳技术研究东亚飞蝗、大垫尖翅蝗、中华稻蝗、中华蚱蜢、亚洲小车蝗和黄胫小车蝗6个种的种群遗传结构及其分化.在所检测的13个基因座位(Ak、Ck、G3pd、Gpi、Hk-1、Hk-2、Idh-1、Idh-2、Ldh、Mdh-1、Mdh-2、Mdph、Pgm)中,有2个基因座位(G3pdh和Mdh-2)在6个种群中均为单态(0.95标准);5个基因座位(Gpi、Hk-1、Hk-2、Mdph和Pgm)在6个种群中均为多态;其余的基因座位至少在一个种群内有两个以上的等位基因.除Gpi和Mdh-1在多数种群符合哈-温(H-W)平衡期望值,其余大多数基因座位的基因型频率显著偏离哈-温(H-W)平衡(P<0.05).从A、P、Ho和He值可知,中华蚱蜢的遗传多样性最低,其次是亚洲小车蝗和黄胫小车蝗,而大垫尖翅蝗、东亚飞蝗和中华稻蝗遗传多样性均较高.根据Roger's遗传距离用非加权算术平均法(UPGMA)进行的聚类分析符合传统形态学和细胞学研究结果.     

条斑星鲽连续性鳍细胞系的建立与鉴定

为了建立条斑星鲽鳍细胞系,为其细胞工程及病毒学研究奠定基础,对经Ⅱ型胶原酶和透明质酸酶联合消化法所得鳍组织碎块分别用DMEM/F12、L-15和M199培养液(pH7.2)在18～26℃进行了鳍组织的体外培养,并在所筛选出的最适培养液和培养温度下通过添加羧甲基壳寡糖、碱性成纤维样生长因子(bFGF)和I型胰岛素样生长因子(IGF-I)启动了鳍细胞的原代培养。体外培养结果显示,条斑星鲽鳍细胞的最适培养液为DMEM/F12培养液,最适培养温度为22℃,原代培养鳍细胞的生长分裂状态旺盛,细胞形态主要为成纤维样,20d后便可形成汇合细胞单层,经过连续的继代培养已建立了条斑星鲽的连续性鳍细胞系,目前已传至第135代;生长特性检测结果显示,第60代鳍细胞系细胞的群体倍增时间为56.9h,其生长分裂状态依然十分旺盛;染色体分析结果显示,第60代鳍细胞系细胞虽然出现了染色体的非整倍性,但其特征性染色体数目仍为46条,并具有2sm+44t的正常二倍体核型,证明所建立的细胞系确为条斑星鲽连续性鳍细胞系。该细胞系为条斑星鲽的细胞工程育种和病毒-细胞相互作用提供了一个理想的体外研究体系,具有重要的理论意义和应用价值。     

基于EST-SSR分子标记的广西海伦兜兰遗传多样性研究

海伦兜兰（Paphiopedilum helenae）具有极高的观赏价值,为国家一级重点保护野生植物。本研究利用EST-SSR分子标记技术对广西海伦兜兰的5个居群（71个样品）的遗传多样性和遗传结构进行分析,旨在为该种质资源的有效保护与利用提供参考依据。结果表明：海伦兜兰具有中等遗传多样性,多态位点百分比（PPB）为90%-100%,期望杂合度（H_e）平均值为0.451,香农信息指数（I）平均值为0.801。海伦兜兰绝大多数（93%）遗传变异存在于居群内,仅有7%遗传变异存在于居群间,居群内的遗传变异大于居群间的变异,基因流（N_m）平均值为4.916。非加权组平均法（UPGMA）、主坐标分析（PCoA）和Structure分析结果表明,聚类并未严格按照地理位置划分,广西邦亮长臂猿国家级自然保护区靖西市（JX）和龙州县下冻镇春秀村（LZ）居群的遗传多样性较高,应作为海伦兜兰重点保护单元进行保护。     

玫瑰冠鸡资源群的微卫星多态性分析

利用8个微卫星DNA标记分析了玫瑰冠鸡(50只)及其杂交鸡(40只)的群体遗传变异。计算了各群体在各位点上的等位基因频率,并据此计算出各群体的平均遗传杂合度、多态信息含量和有效等位基因数。结果表明:2个鸡群在8个微卫星座位上的基因频率存在明显的差异。所选的8个微卫星座位均为高度多态,可作为有效的遗传标记用于鸡群体遗传多样性的分析。     

QTL analysis of the rice seedling cold tolerance in a double haploid population derived from anther culture of a hybrid betweenindica andjaponica rice

A doubled haploid population, derived from anther culture of F₁ hybrid between a typicalindica cv. and ajaponica cv. has been used to investigate the seedling cold tolerance (SCT) in growth cabinet. By dynamically analyzing every day’s survival percentages of the parents and DH lines under 7-d cold plus 9-d normal temperature condition, the quantitative trait loci (QTLs) for SCT have been mapped based on a molecular linkage map constructed from this population. The results show that two parents had significant differences in SCT and the segregation of SCT in DH lines was basically a continuous distribution with most serious injury on the 6th d of the cold treatment. A total of 4 QTLs for SCT have been identified on chromosomes 1, 2, 3 and 4 respectively. The additive effects of qSCT-1, qSCT-2 and qSCT-3 have been contributed by thejaponica cv JX17, but that of qSCT-4 has been contributed by theindica cv ZYQ8. The mechanism of SCT seems complicated since the above 4 QTLs detected at different stages during the treatment. Further study on the genotypes for these SCT QTLs in the DH lines shows transgressive segregation. It is demonstrated that the lines with stronger SCT over JXI7 have 3–4 loci for SCT. Integration of these QTLs into an appropriate variety may lead to a successful rice breeding program for cold tolerance.     

适用于野生软枣猕猴桃遗传多样性分析的SSR引物筛选

为进一步研究野生软枣猕猴桃种质资源的遗传多样性,以软枣猕猴桃基因组DNA为模板,优化了SSR反应体系并确定了反应条件,筛选出了多态性较高的标记.从来自中华猕猴桃基因组的65对SSR引物初步筛选出15对有较好扩增带型、增稳定性好的引物,其中较好多态性的引物有12对.用筛选的引物进一步对四川天全二郎山的野生软枣猕猴桃群体进行了SSR分析,在15个个体中共检测出了141个等位基因,平均每个位点检测到9.4个等位基因,平均PIC值为0.745.     

中国鲎3个地理群体遗传多样性的等位酶分析

应用等位酶技术对漳浦、连江和温州3个地理群体的中国鲎遗传多态性进行分析.实验结果发现：6个酶系统的10个位点中有6个为多态性位点,共获得21个等位基因;大部分位点在3个群体中偏离Hardy-Weinberg平衡;在物种水平上,有效等位基因数为1.635,观察杂合度为0.456,期望杂合度为0.306,香侬指数为0.48...     

野生和人工选育黄河鲤遗传多样性的ISSR分析

利用ISSR分子标记技术,对采集于长垣天然文岩渠(野生群体)、河南省水产科学研究院(黄河品系Ⅰ群体)、小浪底水库(野生群体)、河南黄河鲤鱼良种场(豫选黄河鲤群体)的4个群体黄河鲤的120个个体的遗传多样性进行了分析.8个ISSR引物共获得64个扩增位点,其中多态位点31个,多态位点比例为48.44%.4个群体的多态位点比例分别为14.06%,32.81%,34.38%和20.31%,遗传距离分别为0.0576、0.1657、0.1674和0.0800,Nei基因多样性分别为0.0538、0.1099、0.1308和0.0723,Shannon信息指数分别为0.0784、0.1669、0.1923和0.1072.文岩渠群体与黄河品系Ⅰ群体遗传距离最远(0.1717),与小浪底水库群体遗传距离最近(0.0558).表明这些群体间发生了较弱的遗传分化.UPGMA聚类结果为文岩渠群体与小浪底群体先聚在一起,其次是与豫选黄河鲤,最后与黄河鲤品系Ⅰ相聚.用AMOVA进行遗传变异方差分析得到遗传变异固定指数(Φst=0.0179,P0.05),显示黄河鲤大部分变异(98.21%)发生在群体内,群体间变异较小(1.79%).说明人工选育群体的遗传结构尚未发生明显变化.     

Subcellular localization and inductive expression of the dsRNA-dependent protein kinase PKR from Japanese flounder, Paralichthys olivaceus

The double-stranded RNA (dsRNA)-dependent protein kinase (PKR) belongs to the eIF2α kinase family and plays a critical role in interferon (IFN)-mediated antiviral response. Recently, in Japanese flounder (Paralichthys olivaceus), a PKR gene has been identified. In this study, we showed that PoPKR localized to the cytoplasm, and the dsRNA-binding motifs (dsRBMs) played a determinative role in protein localization. In cultured FEC cells, PoPKR was detected at a low level of constitutive expression but was highly induced after treatment with UV-inactivated grass carp hemorrhagic virus, active SMRV and Poly I:C although with different expression kinetics. In flounder, PoPKR was ubiquitously distributed in all tested tissues, and SMRV infection resulted in significant upregulation at mRNA and protein levels. In order to reveal the role of PoPKR in host antiviral response, its expression upon exposure to various inducers was characterized and further compared with that of PoHRI, which is another eIF2α kinase of flounder. Interestingly, expression comparison revealed that all inducers stimulated upregulation of PoHRI in cultured flounder embryonic cells and fish, with a similar kinetics to PoPKR but to a less extent. These results suggest that, during antiviral immune response, both flounder eIF2α kinases might play similar roles and that PoPKR is the predominant kinase.     

High genetic diversity in population of <Emphasis Type="Italic">Lepturichthys fimbriata</Emphasis> from the Yangtze River revealed by microsatellite DNA analysis

Lepturichthys fimbriata (Günther) is one of the benthic and rock-attached fish species that is typically found in torrential flows of the upper reaches of the Yangtze River in China. Several dams in the Yangtze River (the Ertan Dam, the Three Gorges Dam, the Gezhouba Dam, the Xiluodu Dam and the Xiangjiaba Dam (the latter two dams are under construction)) may have significant effects on the habitat and spawning behaviors of L. fimbriata, and could ultimately threaten the survival of this fish. We studied the population genetic diversity of L. fimbriata samples collected at three sites within the Yangtze River and one of its tributaries, the Yalong River. Genetic diversity patterns were determined by analyzing genetic data from 14 polymorphic microsatellite loci isolated in Jinshaia sinensis (Sauvage et Dabry). A high genetic diversity among these L. fimbriata local populations was indicat- ed by the number of microsatellite alleles (A) and the expected heterozygosity. No reductions of genetic diversity in any L. fimbriata population were observed. However, significant population differentiations were observed among three local populations by pairwise comparisons (P<0.001). We deduced that L. fimbriata local populations were not small ones. In addition, the habitat behaviors of rock-attachment and possible residence of L. fimbriata could account for the genetic differences found in local populations.     

Estimating genetic diversity and sampling strategy for a wild soybean （Glycine soja） population based on different molecular markers

With the rapid development of the global economy and continued increase in world population, natural environments face serious deterioration and change, which has led to the extinction or severe endangerment of many plant species including important crops…     

基于CR及Cytb基因序列的大泷六线鱼野生群体遗传多样性分析

为进一步了解山东省内大泷六线鱼(Hexagrammos otakii)群体遗传背景和分化情况,合理保护与开发利用渔业资源,选取线粒体控制区(CR)和细胞色素b基因(Cytb)对分布于山东省近海沿岸的3个大泷六线鱼野生群体以及北黄海1个离岸大泷六线鱼野生群体的线粒体基因序列进行对比,分析它们的序列特征、遗传多样性和种群历史动态情况。经PCR扩增得到大泷六线鱼野生群体的CR和Cytb基因序列,全长分别为485bp和651bp。基于CR基因序列共检测到40个多态位点,单倍型多样性平均值为0.908,核苷酸多样性平均值为0.006,定义了53种单倍型。基于Cytb基因序列检测到56个多态位点,转换颠换比值为19.04,种内单倍型多样性平均值为0.934,核苷酸多样性平均值为0.005,定义了38种单倍型。对比分析表明,4个大泷六线鱼野生群体均具有高单倍型多样性和低核苷酸多样性。NJ系统进化树、群体间/内平均遗传距离、Fst值、基因流及AMOVA分析结果显示:山东省内近海沿岸大泷六线鱼野生群体和北黄海离岸野生群体遗传差异不显著,群体间存在频繁的基因交流,未形成显著的遗传分化。种群动态结果表明大泷六线鱼于更新世晚期经历了快速扩张,并形成了现有遗传格局。山东省内近海沿岸大泷六线鱼野生群体和北黄海离岸野生群体遗传结构之间不存在显著的地理隔离,这可能与人为增殖放流和秋冬季节黄海沿岸流及暖流有关,从而使得群体之间基因交流广泛。     

Efficient indica and japonica rice identification based on the InDel molecular method: Its implication in rice breeding and evolutionary research

An efficient molecular method for the accurate and efficient identification of indica and japonica rice was created based on the polymorphisms of insertion/deletion (InDel) DNA fragments obtained from the basic local alignment search tool (BLAST) to the entire genomic sequences of indica (93-11) and japonica rice (Nipponbare). The 45 InDel loci were validated experimentally by the polymerase chain reaction (PCR) and polyacrylamide gel electrophoresis (PAGE) in 44 typical indica and japonica rice varieties, including 93-11 and Nipponbare. A neutrality test of the data matrix generated from electrophoretic banding patterns of various InDel loci indicated that 34 InDel loci were strongly associated with the differentiation of indica and japonica rice. More extensive analyses involving cultivated rice varieties from 11 Asian countries, and 12 wild Oryza species with various origins confirmed that indica and japonica characteristics could accurately be determined via calculating the average frequency of indica- or japonica-specific alleles on different InDel loci across the rice genome. This method was named as the “InDel molecular index” that combines molecular and statistical methods in determining the indica and japonica characteristics of rice varieties. Compared with the traditional methods based essentially on morphology, the InDel molecular index provides a very accurate, rapid, simple, and efficient method for identifying indica and japonica rice. In addition, the InDel index can be used to determine indica or japonica characteristics of wild Oryza species, which largely extends the utility of this method. The InDel molecular index provides a new tool for the effective selection of appropriate indica or japonica rice germplasm in rice breeding. It also offers a novel model for the study of the origin, evolution, and genetic differentiation of indica and japonica rice adapted to various environmental changes.