突变型大肠杆菌二氢叶酸还原酶基因的合成

利用PCR定点突变技术，以野生型大肠杆菌二氢叶酸还原酶基因为模板，获取3种突变型(Cys85Ser．Cysl51Ser，Cys85／151Ser)二氢叶酸还原酶(DHFR)基因．用限制性内切酶BamH Ⅰ与Pst Ⅰ将3种突变基因片段插入到克隆载体pUC18上，进行蓝白筛选，将筛选的克隆进行DNA序列测定．     

Statistical analysis on adaptive evolution of SQUA genes in angiosperms

SQUAMOSA (SQUA) subfamily includes important perianth identity genes of MADS-box gene family. SQUA genes of Dendrocalamus latiflorus were sequenced, and phylogenetic form on SQUA genes in angiosperms was analyzed. Relative rate and adaptive evolution after SQUA gene duplication in recent common ancestor of monocots and eudicots were analyzed using the methods of relative rate test, statistic on synonymous and non-synonymous coden substitution sites and likelihood rate test. The results show that both of relative rate and synonymous and non-synonymous coden substitution in eudicot clade are significantly higher than those in monocot clade, and the value of dN/ds uncovered possible positive selective pressure in eudicot clade.     

Genetic analysis and mapping of rice （Oryza sativa L.） male-sterile （OsMS-L） mutant

A rice male-sterile mutant OsMS-L of japonica cultivar 9522 background, was obtained in M4 population treated with ^60Co γ-Ray. Genetic analysis indicated that the male.sterile phenotype was controlled by a single recessive gene. Results of tissue section showed that at microspore stage, OsMS-L tapetum was retarded. Then tapetal calls expanded and microspores degenerated. No matured pollens were observed in OsMS-L anther locus. To map OsMS-L locus, an F2 population was constructed from the cross between the OsMS-L (japonica) and LongTeFu B(indica). Firstly, the OsMS-L locus was roughly mapped between two SSR markers, RM109 and RM7562 on chromosome 2. And then eleven polymorphic markers were developed for further fine fine-mapping. At last the OsMS-L locus was mapped between the two lnDel markers, Lhsl0 and Lhs6 with genetic distance of 0.4 cM, respectively. The region was delimited to 133 kb. All these results were useful for further cloning and functional analysis of OsMS-L.     

Assessment on alien herbicide-resistant gene flow among crucifers by sexual compatibility

Cross-pollination and high cross-compatibility are frequently found in many cruciferous plants, which implies that there might be potential risk of alien gene flow among crucifers. In this work, the alien glyphosate-resistant gene flow between GM （Genetically modified） rapeseed variety Q3 （Brassica napus） and 6 varieties from 5 cruciferous species was assessed by sexual compatibility through aniline blue fluorescence observation, manual cross, mentor pollination with the mixture pollen and DNA identification. The results indicated that the compatibility indices of two B. napus varieties, B. juncea vat. multiceps Tsen et Lee and Chinese cabbage （B. chinensis） （as female） with Q3 （as male） were 16.15, 12.77, 2.345 and 0.85, respectively. FI plants were examined by PCR and all of these samples were confirmed as positive. Crossing between Orychophragmus violaceus, Descurainia sophia （as female） and Q3 （as male） expressed high cross-incompatibility. Fluorescence microscope observation indicated that growing of pollen tube terminated at the upper 1/3 part of the style of O. violaceus and on the stigma surface of D. sophia. The mentor plants were further analyzed by PCR and all were shown to be negative. Under the neighboring growth and natural pollination conditions, the rates of gene flow to two B. napus varieties were 2.3278% and 2.1487%, respectively, B. juncea var. multiceps Tsen et Lee and Chinese cabbage were 1.0157% and 0.9236%, respectively. No gene flow was observed in O. violaceus and D. sophia. Frequency of gene flow was highly correlated with sexual compatibility. Due to possible alien gene flow among crucifers, special consideration and care should be taken to grow GM rapeseed.     

Neurobiological disruptions induced in brains of the rats fed with mercury contaminated rice collected from experimental fields in Guizhou Province, China

Guizhou Province, located in the southwestern China, is an important mercury （Hg） production center. High levels of Hg were found in local environmental and biomass samples. The neurobiological disruptions induced in brains of rats fed with Hg polluted rice collected from two experimental plots in Guizhou Province were studied. The plots are close to the Hg mine of Wanshang area and a chemical plant （where Hg is used as a catalyst for the production of acetaldehyde） of Qingzhen area. Rats were fed for 30 d before toxicological end points were tested. The results showed that the accumulations of Hg in rat＇s brain of Qingzhen chemical plant group and Wanshang mine group were significantly higher than those of Shanghai group （control group） （P〈0.01）. The neural transmitters, including acetylcholine （Ach）, acetylcholine enzyme （AchE）, nitric oxide and nitricoxide synthase （NOS）, in rat＇s brain were significantly affected. Oxidative stress was observed in the exposure groups. The expression levels of the immediately early gene （lEG）, c-fos and c-FOS proteins were induced by the Hg polluted rice. It was noted that the lEG （c-fos） participated in the neurotoxic process induced by the Hg polluted rice. lEG response to Hg might be dependent on interactions between neural transmitter and oxidative damage, c.fos could be used as an effective index of detecting and assessing the neurotoxicity of Hg. The results suggested that it should be alert to mental health problem in human beings eating Hg-polluted food. More efforts are necessary to protect the local ecosystem and human health in the Hg polluted areas such as Wangshan and Qingzhen in Guizhou Province of China.     

Fine mapping of an incomplete recessive gene for leaf rolling in rice （Oryza sativa L.）

Genetic analysis and fine mapping of genes controlling leaf rolling were conducted using two backcrossed generations （BC4F2, BC4F3） derived from a cross between QMX, a non-rolled leaf cultivar as a recurrent parent, and JZB, a rolled leaf NIL of ZB as a donor parent. Results indicated that leaf rolling was mainly controlled by an incompletely recessive major gene, namely rl（t）, and at the same time, affected by quantitative trait loci （QTLs） and/or the environment. A genetic linkage map was constructed using MAPMAKER/EXP3.0 with eight polymorphic markers on chromosome 2, which were screened by BAS method from 500 SSR markers and 15 newly developed insertion/deletion （InDel） markers. The position of rl（t） was estimated with composite interval mapping （CIM） method using WinQTLcart2.5. Gene rl（t） was mapped between markers InDel 112 and RM3763, and 1.0 cM away from InDel 112 using 241 plants in BC4F2 population. To fine map r（t）, one BC4F3 line with 855 plants was generated from one semi-rolled leaf plant in BC4F2. Four new polymorphic InDel markers were developed, including InDel 112.6 and InDel 113 located between markers InDe1112 and RM3763. Based on the information of recombination offered by 191 rolled leaf plants and 185 non-rolled leaf plants from the BC4F3 line ,we mapped r（t） to a 137-kb region between markers InDel 112.6 and InDel 113. Homologous gene analysis suggested that r（t）was probably related to the process of leaf development regulated by microRNA.     

肌肉抑制素基因与RNA干扰的应用前景

本文综述了肌肉抑制素基因的研究进展,以及RNA干扰技术的功能.展望了RNA干扰技术在肌肉抑制素基因中的应用前景.     

生物芯片技术的研究进展

生物芯片技术是近些年来发展迅速的一项高新技术.论述了生物芯片的分类、应用及研究进展,并对生物芯片的发展前景作了展望.     

河套蜜瓜ACC氧化酶基因cDNA部分片段的克隆和序列分析

以河套蜜瓜(Cucumis melo L．ev Hetao)成熟果实为材料，用硫氰酸胍法提取总RNA，以oligo(dT)15为引物，反转录合成cDNA，利用一对甜瓜ACC氧化酶(1-aminocyclopropane-1-earboxylate oxidase，ACO)基因cDNA的特异性引物进行聚合酶链式反应(PCR)扩增。获得了预期大小的cDNA片段．将此片段克隆于pUCl9质粒中，进行DNA序列分析．序列分析结果表明该片段为545bp，编码甜瓜ACO的第126至第306个氨基酸．该河套蜜瓜ACO基因cDNA序列与Andes甜瓜、Cantaloup charentais甜瓜和哈密瓜的相应序列进行比较，其核苷酸序列与Andes甜瓜的同源性为99．4％；与Cantaloup charentais甜瓜和哈密瓜的同源性均为99．6％。与其对应的氨基酸序列的同源性均为99．4％．     

Si-1基因中一个多态位点的初步分析

 通过PCR-RFLP技术分析了Si-1基因的外显子在肿瘤样本与正常人群外周血液样本的突变情况,初步确定了Si-1基因的第15号外显子的第56位发生点突变,突变类型为C—T的置换突变,其编码的P(脯氨酸)变化为S(丝氨酸).在正常人群外周血液样本中的突变频率为14%,而在肿瘤中的突变频率为24.7%,尤其在肠癌中的突变频率高达51.5%.上述结果表明Si-1基因的外显子的突变与肿瘤间有一定关系.