|本期目录/Table of Contents|

[1]张武,田润,申佩弘,等.费氏中华根瘤菌15142的cysDN基因克隆及其相关功能[J].应用与环境生物学报,2011,17(06):864-868.[doi:10.3724/SP.J.1145.2011.00864]
 ZHANG Wu,TIAN Run,SHEN Peihong,et al.Cloning and Related Function of cysDN in Sinorhizobium fredii 15142[J].Chinese Journal of Applied & Environmental Biology,2011,17(06):864-868.[doi:10.3724/SP.J.1145.2011.00864]
点击复制

费氏中华根瘤菌15142的cysDN基因克隆及其相关功能()
分享到:

《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
17卷
期数:
2011年06期
页码:
864-868
栏目:
研究论文
出版日期:
2011-12-25

文章信息/Info

Title:
Cloning and Related Function of cysDN in Sinorhizobium fredii 15142
作者:
张武田润申佩弘蒋承建许兢李俊芳武波
(1广西大学生命科学与技术学院 南宁 530005)
(2微生物及植物遗传工程教育部重点实验室 南宁 530005)
(3广西亚热带生物资源保护利用重点实验室 南宁 530005)
(4广西民族大学化学与生态工程学院 南宁 530007)
Author(s):
ZHANG WuTIAN RunSHEN PeihongJIANG ChengjianXU JingLI JunfangWU Bo
(1College of Life Science and Technology, Guangxi University, Nanning 530005, China)
(2Key Laboratory of Ministry of Education for Microbial and Plant Genetic Engineering, Nanning 530005, China)
(3Guangxi Key Laboratory of Subtropical Bioresource Conservation and Utilization, Nanning 530005, China)
(4College of Chemistry and Ecology Engineering, Guangxi University for Nationalities, Nanning 530007, China)
关键词:
费氏中华根瘤菌硫代谢cysDN基因克隆同源单交换植株试验
Keywords:
Sinorhizobium fredii sulfate assimilation cysDN gene clone homologous single crossover plant test
分类号:
Q939.114 : Q78
DOI:
10.3724/SP.J.1145.2011.00864
文献标志码:
A
摘要:
为确定cysDN操纵子的功能及对根瘤菌结瘤的影响,通过三亲本接合,将质粒转座子pTnMod-RKm′随机插入费氏中华根瘤菌15142中,建成随机插入突变体库,随后通过含有不同硫源的MM培养基的筛选得到一株不能利用硫酸盐但能够利用半胱氨酸的突变体. 进一步克隆和测序分析后发现该操纵子与已报道的Sinorhizobium sp. strain BR816的cysDN在核苷酸水平上有92%的相似性,在氨基酸水平上有96%的相似性. 用自杀质粒pK18mob分别构建含有cysD部分片段和cysN部分片段的重组质粒,通过三亲本接合导入出发菌株15142中,经过同源单交换,分别获得cysD的pK18mob正反向插入突变株cysDF/15142 以及cysDR/15142和cysN的pK18mob正反向插入突变株cysNF/15142与cysNR/15142. 用广谱宿主质粒pLAFRJ载体连接完整操纵子cysDN构建互补质粒cysDN+pLAFRJ,将该质粒通过三亲本接合导入突变株中,获得互补菌株. 用不同硫源的液体MM培养基培养,发现互补菌株能够补回突变菌株不能利用硫酸盐作为唯一硫源的缺陷,说明cysDN操纵子确实与硫酸盐同化途径有关;植株试验表明突变株比出发菌株推迟结瘤1~2 d,固氮酶活也比出发菌株稍低;竞争结瘤试验表明突变菌株占瘤率较差,但在平均瘤数、平均瘤重、平均植株干重上则无差异. 图3 表4 参10
Abstract:
In order to know function of cysDN and its effect on plant nodulation, a random insertion mutant library of Sinorhizobium fredii 15142 was constructed by three parental hybridization with the plasmid pTnMod-RKm’ as transposon vector. A mutant strain which could not metabolize sulfate while could use cysteine as sulfur source, was selected by MM medium with different sulfur sources from the library. Molecular cloning and sequence analysis showed that this operon had 92% and 96% similarity with the cysDN gene sequence of Sinorhizobium sp. BR816 at nucleotide and amino acid level, respectively. With the suicide plasmid pK18mob, the polar mutant cysDF/15142, cysNF/15142 and non-polar mutant cysDR/15142, cysNR/15142 inactivated in cysD and cysN were constructed through homologous recombination. A recombinant plasmid pLAFRJ+cysDN was constructed by using plasmid pLAFRJ ligated with cysDN, and transferred into above mutants to obtain complement strains. The results of growth tests found that the mutants had no growth on the MM medium with sulfate as sulfur source but the complement strains could grow normally, indicating that cysDN operon was related to sulfate assimilation. The plant tests indicated that the mutant strains delayed 1~2 d to nodulate than the wild strain, and nitrogenase activity and nodulation efficiency in mutants were slightly lower than those of the original strain, but there were no differences in the number and weight of nodules and the dry weight of plant. Fig 3, Tab 4, Ref 10

参考文献/References:

1 Roche P, Debelle F, Maillet F, Lerouge P. Molecular basis of symbiotic host specificity in Rhizobium meliloti: nodH and nodPQ genes encode the sulfation of lipooligosaccharide signals. Cell, 1991, 67 (6): 1131~1143
2 Suiko M, Fernando PH, Sakakibara Y. Post-translational modification of protein by tyrosine sulfation: Active sulfate PAPS is the essential substrate for this modification. Nucleic Acids Symp Ser, 1992, 27 (2): 183~184
3 Sekowska1 A, Kung HF, Danchin A. Sulfur metabolism in Escherichia coli and related bacteria: Facts and fiction. J Mol Microbiol Biotechnol, 2000, 2 (2): 145~177
4 Reuter K, Mofid MR, Marahiel MA. Crystal structure of the surfactin synthetase-activating enzyme Sfp: A prototype of the 4’-phosphopantetheinyltransferase superfamily. EMBO J, 1999, 18 (23): 6823~6831
5 Shen P (沈萍), Fan XR (范秀容), Li GW (李广武). 微生物学实验. Beijing, China: Higher Education Press (北京: 高等教育出版社), 1999. 22l
6 Robertsen BK, Aiman P, Darvill AG. Host-symbiont interactions: V. The structure of acidic extracellular polysaccharides secreted by Rhizobium leguminosarum and Rhizobium trifolli. Plant Physiol, 1981, 67 (3): 389~400
7 Xue YL (薛应龙). 植物生理学实验手册. Shanghai, China: Shanghai Science & Technology Press (上海: 上海科学技术出版社), 1985. 259~264
8 Hussain S, Ali V, Jeelani G, Nozaki T. Isoform-dependent feedback regulation of serine O-acetyltransferase isoenzymes involved in L-cysteine biosynthesis of Entamoeba histolytica. Mol Biochem Parasitol, 2009, 163 (1): 39~47
9 Schnell R, Schneider G. Structural enzymology of sulfur metabolism in Mycobacterium tuberculosis. Biochem Biophys Res Commun, 2010, 396 (1): 33~38
10 Abola AP, Willits MG, Wang RC. Reduction of adenosine-5’-phosphosulfate instead of 3’-phosphoadenosine-5’-phosphosulfate in cysteine biosynthesis by Rhizobium meliloti and other members of the family Rhizobiaceae. J Bacteriol, 1999, 181 (17): 5280~5287

相似文献/References:

[1]宋张杨,马婷婷,周燕,等.不同根瘤菌中突变cysDN基因对硫酸盐同化途径的影响[J].应用与环境生物学报,2015,21(02):242.[doi:10.3724/SP.J.1145.2014.10020]
 SONG Zhangyang,MA Tingting,ZHOU Yan,et al.Effect of cysDN genes in different rhziobium on the sulfate assimilation pathway[J].Chinese Journal of Applied & Environmental Biology,2015,21(06):242.[doi:10.3724/SP.J.1145.2014.10020]
[2]成家龙 陈大松 李友国**.高效固氮大豆根瘤菌SMH12的全基因组测序及比较基因组分析*[J].应用与环境生物学报,2020,26(01):1.[doi:10.19675/j.cnki.1006-687x.2019.04011]
 CHENG Jialong,CHEN Dasong & LI Youguo**.Whole genome sequencing of highly nitrogen-fixing strain Sinorhizobium fredii SMH12 and genome comparative analysis *[J].Chinese Journal of Applied & Environmental Biology,2020,26(06):1.[doi:10.19675/j.cnki.1006-687x.2019.04011]

备注/Memo

备注/Memo:
国家重点基础研究发展计划项目(No. 2010CB126502)资助
更新日期/Last Update: 2011-12-31