水稻AP2/EREBP转录因子基因OsDRE和OsRAF的功能分析

AP2是一个大的转录因子家族，因其成员含有一个60-70 个氨基酸残基的保守结构域即AP2结构域而得名，它们的功能涉及植物花的发育以及植物对生物或非生物逆境的应答反应。根据它们所含的AP2 结构域的数目，这个家族可分为AP2亚家族和EREBP 亚家族。 AP2亚家族成员含两个AP2结构域，EREBP 亚家族成员含一个AP2结构域。一般来说，AP2 亚家族的成员主要参与植物发育过程的调控;EREBP 亚家族成员则主要参与对逆境的应答反应。按照它们响应外界刺激的类型和AP2结构域中结合顺式作用元件的核心氨基酸的不同，EREBP亚家族又可分为ERF和CBF/DREB两大类群，ERF 类主要响应生物类逆境的诱导，CBF/DREB类则响应干旱、低温等非生物胁迫的刺激。 根据AP2保守结构域搜索，水稻基因组中一共有147个AP2/EREBP成员，但其中功能得到证实的还非常有限。为了解更多AP2基因在植物生长发育过程中的功能，我们先从水稻基因组数据库中搜索到含有AP2/EREBP 结构域的推测基因序列，选择其中40个扩增并成功扩增出31个，将这些DNA片段点在尼龙膜上，然后用水稻叶片cDNA 作模板标记探针，与固定在膜上的推测基因杂交。杂交结果作为选择基因进行功能分析的重要依据。 OsDRE就是我们选择进行研究的一个表达较强的基因。首先，通过RACE 克隆得到OsDRE 的cDNA全长 1589bp，它编码318个氨基酸。Blast 搜索和保守结构域序列比对分析以及进化树分析显示它是一个新的ERF 基因。RT-PCR分析表明该基因在水稻各种组织中表达量比较一致，而且，OsDRE 既不对植物生长物质如乙烯，水杨酸（SA），茉莉酸甲酯（MJ），脱落酸（ABA），赤霉素（GA3），油菜素内酯（BR）的诱导起反应，同样也不响应环境因子如低温、干旱条件的处理。这些结果说明OsDRE是一个不响应胁迫相关因素的诱导的、组成型表达的水稻基因。用OsDRE的非保守区域构建的RNAi 载体转入水稻后未能使转基因水稻产生异常表型，然而，OsDRE在水稻和拟南芥中的过量表达都导致转基因植物出现植株矮小、开花延迟、生长周期延长以及育性降低等表型，说明OsDRE对生长和发育的影响在水稻和拟南芥中是一致的。 基于以上原因，我们选择在遗传分析方面有明显优势的拟南芥作为材料，对OsDRE基因功能进行研究并得出以下结论：（1）瞬时表达和随后的过量表达证明OsDRE定位于细胞核中，过量表达OsDRE引起转基因植株的生长周期变长、抽苔时间延迟和抽苔时莲座叶的数量增多;（2）过量表达OsDRE通过一种不影响细胞数量的方式抑制了细胞的膨胀从而导致植物器官以致整个植株变小，而且，在此过程中部分器官的形态也受到了影响;（3）OsDRE过量表达能激活已知位于乙烯信号途径下游的基因表达并且转基因植株幼苗在黑暗中出现下胚轴及根缩短变粗的现象，提示OsDRE 可能部分参与了乙烯信号途径下游的反应。 除此之外，我们还初步分析了另一个EREBP基因，并将其命名为OsRAF。氨基酸序列分析表明该基因与大麦的RAF 基因在蛋白水平上相似性最高。Northern 杂交结果进一步显示，与RAF 一样，OsRAF 也是根中优势表达的基因，并且它的表达量在乙烯或低温的诱导下增加。对转基因植株的观察和瞬时表达表明OsRAF 定位于细胞核中。 综上所述，对水稻基因OsDRE 和OsRAF的分析表明， OsDRE是一个新的ERF基因，它不受乙烯等因素的诱导并且过量表达该基因导致转基因植株出现细胞膨胀受到抑制等一系列的表型。另外，OsRAF在水稻根中优势表达并受乙烯和低温的诱导，目前，与之相关的功能研究正在进行。

Construction of a BAC library for Chinese amphioxus Branchiostoma belcheri and identification of clones containing Amphi-pax genes

Amphioxus is a crucial organism for the study of vertebrate evolution. Although a genomic BAC library of Branchiostoma floridae has been constructed, we report here another BAC library construction of its distant relative species Branchiostoma belcheri. The amphioxus BAC library established in present study consists of 45,312 clones arrayed in one hundred and eighteen 384-well plates. The average insert fragment size was 120 kb estimated by Pulsed Field Gel Electrophoresis (PFGE) analysis of 318 randomly selected clones. The representation of the library is about 12 equivalent to the genome, allowing a 99.9995% probability of recovering any specific sequence of interest. We further screened the library with 4 single copied Amphi-Pax genes and identified total of 26 positive clones with average of 6.5 clones for each gene. The result indicates this library is well suited for many applications and should also serve as a useful complemental resource for the scientific community.

一种改进的高分辨率RFLPs 分析

国家自然科学基金和云南省自然科学基金资助

Heavy metal concentration in South Vietnam waters

Data obtained from investigations which had been carried out during 1998-1999 in the waters of South Vietnam indicate that concentrations of Fe and Zn were usually higher than permissible limits. In the East South Vietnam waters, Fe concentration varied from 77 to 4450g/l (mean 1045g/l) and Zn concentration varied from 7.24 to 74.1g/l (mean 26.7g/l), Pollution coefficients of the two metals were 10.4 and 2.7 respectively. In the Gulf of Thailand, variation range of Fe was 133-850g/l (mean 318g/l) and of Zn was 4.8-167.8g/l (mean 23.46g/l); corresponding pollution coefficients were 3.2 and 2.3. Mn, Cu, As had mean concentrations lower than permissible limits. In the East South Vietnam waters, variation range for Mn was 9.0-444.6g/l (mean 57.0g/l), for Cu was 1.0- 20.3g/l (mean 3.1g/l) and for As was 1.0-10.5 g/l (mean 3.7 g/l). In the Gulf of Thailand, Mn concentration varied from 1.2 to 410.5g/l (mean 19.1g/l), Cu concentration varied from 1.2 to 14.7g/l (mean 3.7g/l), variation range of As concentration was 1.0-13.3g/l with the mean value was 3.6g/l. Scarce data relating to Hg, Cd, Pb showed that their concentrations in the East South Vietnam waters were negligible (usually < 1.0g/l).

浙江省兼用型地方鸡种亲缘关系的研究

测定了浙江省三个兼用型品种(萧山鸡、白银耳鸡、灵昆鸡)539bp的线粒体DNA D-环区序列,计算了各品种间的遗传距离,构建了各鸡种的聚类关系图。结果显示,三个鸡种之间的遗传差异较小,其中灵昆鸡和白银耳鸡关系较近,萧山鸡与这两种鸡关系较远。从品种外形、形成历史及DNA水平等方面综合考虑,灵昆鸡的母系来源可能就是白银耳鸡。结果还显示,萧山鸡是遗传上变异较大的类群,具丰富的遗传多样性。

Taxonomic status of the white-head langur (Trachypithecus francoisi leucoscephalus) inferred from allozyme electrophoresis and random amplified polymorphism DNA (RAPD)

Six sample specimens of Trachypithecus francoisi and 3 of T. leucocephalus were analyzed by use of allozyme electrophoresis and random amplified polymorphism DNA (RAPD) in order to clarify the challenged taxonomic status of the white-head langur. Among the 44 loci surveyed, only 1 locus (PGM-2) was found to be polymorphic. Nei's genetic distance was 0.0025. In total, thirty 10-mer arbitrary primers were used for RAPD analysis, of which 22 generated clear bands. Phylogenetic trees were constructed based on genetic distances using neighbor-joining and UPGMA methods. The results show that T. francoisi and T: leucocephalus are not monophyletic. T. francoisi from Guangxi, China and Vietnam could not be clearly distinguished, and they are not divided into 2 clusters. A t-test was performed to evaluate between genetic distances within and between T. leucocephalus and T. francoisi taxa groups. The statistical test shows that the taxa group within T: leucocephalus and T: francoisi does not significantly differ from that between T: leucocephalus and T: francoisi at the 5% level. Our results suggest that the level of genetic differentiation between T, leucocephalus and T. francoisi is relatively low. Recent gene flow might exist between T. francoisi and T. leucocephalus. Combining morphological features, geographical distribution, allozyme data, RAPD data, and mtDNA sequences, we suggest that the white-head langur might be a subspecies of T. francoisi.

Phylogeographic analysis of mtDNA variation in four ethnic populations from Yunnan Province: new data and a reappraisal

Two sets of mitochondrial DNA (mtDNA) hypervariable segment I (HVS-I) data from four ethnic populations (Tibetan, Va, Dai, and Lahu) from Yunnan Province, China, were analyzed here by using phylogeographic methods. The results suggest that more attention

Comparison of quantum chemistry calculation methods of nucleic acid base-quartets

Nine base-quartets were calculated by six semi-empirical methods and ab initio Hartree-Fork method using STO-3G basis set. The results showed that PM3 method can be use to calculate base quartets, the results of PM3 calculations are close to the ab initio

Impacts of environmental conditions on macrobenthic distribution along the Suez Gulf, Egypt

Increases in coastal development and human activities leading to sedimentation degrade the quality of water; disturb the natural structure and functions of aquatic communities. The Suez Gulf is a large semi-closed area (~625 km long coastline).The assemblages of bottom fauna were studied qualitatively and quantitatively in the shallow intertidal waters along the western coast of the Suez Gulf. The quality of seawater and sediment structures were analyzed. The distribution of macro-benthos included a total of 38 species of Gastropoda and 9 Bivalvia; and 25 species from the other invertebrates included 7 groups namely, Rhizostoma, Polychaeta, Cirripedia, Amphipoda, Isopoda, Decapoda and Echinodermata. The most dominant group among invertebrate groups was the Polychaeta which included 4 species: Hydroides elegans, Perinereis cultilifera, Perinereis nuntia and Ophelina acuminata. The Cirripedia were represented by 3 species namely, Balanus amphitrite, Chithamalus challengeri and Tetraclita squamosa. The variations in the numerical abundance and biomass of bottom fauna studied between the observation periods and at sampling sites. There was a marked increase in benthos biomass at St. IV (Ras Gharib) yielding an average of 318.8 g/m² in which the gastropod community represented the dominant species in collected samples reaching 270.28 g/m² (84.4% of the total biomass) and numerically numbered 116 ind./m². Veliger larvae of bivalves and gastropods appeared to be present in the plankton for long periods and their production seems to be continuous throughout the year. In the intertidal zone of the Suez Gulf, the values of pH varied within narrow limits. Water temperature and salinity seemed to be important in the distribution and abundance of the macro-benthos communities in the study areas. The organic content in shallow intertidal waters and sediments indicated high values in the central part of the Gulf of Suez.

Mercury bioaccumulation in pike (Esox lucius) food chain from Anzali Lagoon, Iran

A number of wide-ranging monitoring studies have been performed in order to estimate the degree of mercury (Hg) contamination in freshwater ecosystems. Knowledge regarding contamination of different levels of the food chain is necessary for estimation of total pollutant input fluxes and subsequent partitioning among different phases in the aquatic system. The growing international concern about this environmental data is closely related to the strongly developing ecological risk assessment activities. In addition,freshwater monitoring outputs hold a key position in the estimation of the Hg dose consumed by the human population as it is highly dependent on fish consumption. So monitoring of Hg in the tissue of edible fish is extremely important because of contaminated fish has caused serious neurological damage to new born babies and adults. Mercury tends to accumulate in fish tissue, particularly, in the form of methyl mercury, which is about 10 times more toxic than inorganic mercury. The Anzali lagoon is one of the biggest wetland of Guilan province, which joins to the Caspian sea. Many Chemical and industrial factories plus agricultural runoffs and urban and rural sewages are major polluting sources of the Anzali wetland. Since many of those polluting sources drain their wastes directly or indirectly into the Anzali wetland and their sewages may be polluted with Hg, this study was conducted to find out the bioaccumulation of Hg bioaccumulation in pike (Esox lucius) food chain from Anzali lagoon, Iran. Sampling were carried out from July 2004 to July 2005, in addition 318 speciments of 9 fish species were collected. T-Hg was measured by LECO AMA 254 Advanced Mercury Analyzer (USA) according to ASTM standard No D-6722. Each sample was analyzed 3 times. Accuracy of T-Hg analysis was checked by running three samples of Standard Reference Materials; SRM 1633b, SRM 2711 & Sra 2709. Detection limit was 0.001 mg/kg in dry weight. The Accuracy degree of analyzor equipment with RSD<%0.05 (N=7) was between %95.5 and %105. In overal eigth fish species were distingushed in the gut content of 87 speciments of pike with age 1-5 year and maximum length 550mm. The max. and min. concentration of T-Hg in dorsal muscle of pjke was 0.2ppm in one year and 1.2ppm in five year class. The mean of T-Hg significantly increased with age and length increased (P<0.05).Mercury accumulation pattern in pike was as well as muscle > liver > spleen (P<0.05). THg content in female was higher than male(P<0.05). In contrast the mean of THg concentration in dorsal muscle of eigth fish species as prey was 0.282, 0.261, 0.328, 0.254, 0.256, 0.286, 0.322 and 0.241 ppm for Carassius auratus gibelio, Hemiculter leucisculus, Blicca bjoerkna transcaucasica, Chalcalburnus mossulensis, Rhodeus sericeus amarus, Gambusia holbrooki, Alburnus charusini hohenackeri & Scardinius Erythrophthalmus respectively.Liner regresion indicated that high degree of relationship between age of pike and Uptak/Intake ratio (R2=%99.12) and indicated that the mercury bioaccumulation in the pike dorsal muscle increased with age increased. BFA was >1 and and indicating the mercury biomagnification in the pike food chain. Trophy level of pike in the Anzali lagoon was estimated as well as 3.5 and 4 . It is generally agreed that Hg concentration in carnivorous fish are higher than in noncarnivorous species.