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Duplication and expression analysis of multicopy miRNA gene family members in Arabidopsis and rice

Cell research 2006年第5期16卷 507-518页

作者： Danhua Jiang~(1,2) Changsong Yin~1 Aiping Yu~(1,3) Xiaofan Zhou~1 Wanqi Liang~(1,4) Zheng Yuan~1 Yun Xu~1 Qingbo Yu~1 Tieqiao Wen~2 Dabing Zhang~(1,4) ~1SJTU-SIBS-PSU Joint center for Life sciences,School of Life Science and biotechnology,shanghai Jiao Tong University 800 Dongchuan road,shanghai 200240, china ~2School of Life Science,shanghai University,99 Shangda road,shanghai 200444,china ~3College of Life and Environment sciences,shanghai Normal University,100 Guilin road, shanghai 200234,china ~4key laboratory of agricultural genetics and breeding,agro-biotech research center,shanghai academy of agricultural sciences,2.01 beidi road,shanghai 2.1106,china SJTU-SIBS-PSUJointcenterforLifesciences SchoolofLifeScienceandBiotechnologyShanghaiJiaoTongUniversity800DongchuanRoadShanghai200240China SchoolofLifeScience ShanghaiUniversity99ShangdaRoadShanghai200444

To understand the expansion of multicopy microRNA (miRNA) families in plants,we localized the reported miRNAgenes from Arabidopsis and rice to their chromosomes,respectively,and observed that 37% of 117 miRNA genes fromArabidopsis and 35% of 173 miRNA genes from rice were segmental duplications in the *** order to characterizewhether the expression diversification has occurred among plant multicopy miRNA family members,we designed PCRprimers targeting 48 predicted miRNA precursors from 10 families in Arabidopsis and *** from RT-PCR datasuggest that the transcribed precursors of members within the same miRNA family were present at different *** addition,although miR160 and miR162 sequences were conserved in Arabidopsis and rice,we found that theexpression patterns of these genes differed between the two *** data suggested that expression diversifica-tion has occurred in multicopy miRNA families,increasing our understanding of the expression regulation of miRNAsin plants.

关键词： gene duplication microRNA multicopy

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Screening and construct specific detection methods of transgenic Huafan No.1 tomato by conventional and real-time PCR

Screening and construct specific detection methods of transg...

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中国植物生理学会第九次全国会议

作者： LitaoYang Huifeng Shen Jianxiu Chen Dabing Zhang ( key laboratory of agricultural genetics and breeding, agro-biotech research center, shanghai academy of agricultural sciences, shanghai 2.1106,) ( School of life Science and biotechnology, shanghai Jiao Tong University, shanghai 200240)

＜正＞ Abstract: Genetically modified （GM） tomatoes have been approved for commercialization in many countries since the first GM tomato FLAVER AVAR permitted for plant in 1994. Also, GM tomato Huafan No.1 with a character of long shelf-life was the first GM plant approved for commercialization in 1996 in china. To meet the requirement of GM tomatoes labeling policy actualization in china since 2.01, the screening and construct specific PCR detection methods for detecting the universal elements transformed into tomato, such as Cauliflower mosaic virus 35S （CaMV35s） promoter and the nopaline synthase （NOS） terminator of agrobacterium tumefaciens, and the specific inserted heterologous DNA sequence between CaMV35s promoter and anti-sense ethylene-forming enzyme （EFE） gene were set up, respectively. To make the detection methods normative, a novel single copy tomato gene LAT52 was also used as an endogenous reference gene in the PCR detection systems. The limit of detection （LOD） of screening and construct specific detection methods for Huafan No.1 was 68 haploid genome copies in conventional PCR detection, and 3 copies in TaqMan real-time PCR detection. The limit of quantitation （LOQ） of screening quantitative PCR assays for Huafan No. 1 was 3 copies and 25 copies for construct specific quantitative PCR,respectively. Two samples with known Huafan No.1 tomato contents were detected using the established conventional and real-time PCR systems, and these results also indicated that the established Huafan No.1 screening and construct specific PCR detection systems were reliable, sensitive, and accurate.

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Cross-breeding and Selection of Flammulina velutipes Hybrid Strains for Industrial Cultivation

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食用菌学报 2008年第4期 31-34页

作者： XU Zhen SHANG Xiaodong SU Rong TAN Qi(key laboratory of Applied Mycological Resources and Utilization,Ministry of Agriculture,the People's Republic of china shanghai research center for Edible Fungi biotechnology and Engineering key laboratory of agricultural genetics and breeding of shanghai Institute of Edible Fungi,shanghai academy of agricultural sciences,shanghai 2.1106,china)

White Flammulina velutipes strains FM0 F21,F10 and F3-W were selected as the parental strains in a single-spore cross-breeding *** basidiospore-derived monokaryons were selected randomly from each parental strain,and mating was carried out according to the mating groups:FM×F21,FM×F3-W and F3-W×*** analysis revealed that hybrid progenies from the FM×F21 mating group exhibited the highest mating rate (79%).However,only 59.2% of these hybrids produced measurable fruit body yields compared with 82.6% and 84.4% of the hybrids generated from FM×F3-W and F10×F3-W crosses respectively,perhaps indicating that certain hybrid progeny were better adapted to the standard set of cultivation conditions adopted in the *** rates for FM×F3-W and F10 x F3-W crosses were 28% and 42%,respectively indicating that the number of mating type genes may vary among the monokaryons constituting each of the three mating *** growth rates of most FM×F21 hybrid progeny ranged between 4.0～7.0 mm/*** values for progeny from FM×F3-W and F10×F3-W crosses were 3.0～6.0 mm/d and 2.0～6.0 mm/d,respectively although a small number of dikaryons from these two mating groups exhibited much higher/lower growth *** correlation between mycelial growth rate and fruit body yield was evident for any of the three sets of hybrid *** bodies produced by all the hybrid progeny were white in color.

关键词： Flammulina velutipes single-spore cross-breeding mycelium growth rate fruit body yield

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Optimization of Electroporation Processing of Bacillus subtilis

Optimization of Electroporation Processing of Bacillus subti...

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2012年环境污染与大众健康学术会议

作者： Jinbin Wang 1,3 , Dachao Chen 1,2 , Ziping Zhu 1 , Ying Xu 1 , Xueming Tang 1,3 1 biotechnology research Institute, shanghai academy of agricultural sciences, 2.01 beidi road, shanghai, 2.1106, china 2 shanghai Ocean University, shanghai, 201306, china 3 key laboratory of agricultural genetics and breeding, shanghai academy of agricultural sciences, 2.01 beidi road, shanghai, 2.1106, china

In order to obtain an optimal transformation efficiency of Bacillus subtilis electroporation experimental program, to optimize the conditions for electroporation of Bacillus subtilis cells, a shuttle vector for Escherichia coli and Bacillus subtilis, pHY300PLK, were used. Factors including growth medium, electroporation medium, recovery Medium, plasmid concentration and pulse strength were investigated. Single factor experiments were employed to optimize the growth medium, electroporation medium, recovery Medium,plasmid concentration and pulse strength. Results showed that the optimal values for concentration of the growth medium electroporation medium, recovery Medium,plasmid concentration and pulse strength are LB+0.5 M sorbitol, 0.5 M sorbitol+0.5 M mannitol+10% Glycero, LB+0.5 M sorbitol+0.38 M mannitol, 20 kV/ cm and 100 ng/μl of Plasmid DNA.

关键词： Bacillus subtilis Electroporation Transformation efficiency

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Genome-wide SNP in silico detection of Lentinula edodes via whole genome sequencing

Genome-wide SNP in silico detection of Lentinula edodes via ...

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中国菌物学会第五届会员代表大会暨2011年学术年会

作者： CHEN Xiang BAO Da-Peng (National research center for Edible Fungi biotechnology and Engineering key laboratory of Applied Mycological Resources and Utilization,Ministry of Agriculture,china shanghai key laboratory of agricultural genetics and breeding Institute of Edible Fungi,shanghai academy of agricultural sciences,shanghai,china)

＜正＞Analysis of single nucleotide polymorphisms（SNPs） emerges as an important tool in evolutionary biology and *** we report the in silico discovery and analysis of SNPs across the whole genome of two protoplast monokaryons（TPMs） of basidiomycete fungi Lentinula edodes via the high-throughput sequencing *** our knowledge,this is a novel study in which different protoplast monokaryons are utilized for SNPs detection and *** sequence information including 210M and 250M nucleotide bases was generated from TPMs yielding approximately 80%of whole genome *** used surpassed 380K and 410K reads of TPMs for the de novo assembly of two contig sets with an overall estimated depth of 5.67-fold and 6.05-fold coverage,*** of the contigs between TPMs identified 108,235 putative SNPs owning a high probability of at least 10-fold *** SNPs reveal TPMs having similar intra-strain SNP frequencies,and the ratio of transition to transversion among these SNPs being similar to the ratio reported in other *** verify the confidence of detected SNPs,25,044 SNPs were mapped onto 43,912 positions that uniformly distributed over the sequenced chromosomes of Coprinopsis *** result shows high sequence conservation among closely related fungal *** addition,the gene structure of TPMs was constructed for SNPs classification according to their distribution in genome ***,this study indicates the utility of using next generation sequencing technologies to identify large numbers of reliable ***,the method combination of SNP detection and novel algorithm allows an efficient approach for the development of high-density genotyping platforms in fungal species.

关键词： Lentinula edodes whole genome sequencing contig assembly SNPs

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Inheritance Patterns of a Strain-specific Sequence Characterized Amplified Region (SCAR) Marker for Lentinula edodes Strain 135

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食用菌学报 2008年第4期 6-10页

作者： ZHANG Meiyan~1 SHANG Xiaodong~1 TAN Qi~(1*) CHEN Mingjie~1 PAN Yingjie~2(~1key laboratory of Applied Mycological Resources and Utilization,Ministry of Agriculture,the People's Republic of china shanghai research center for Edible Fungi biotechnology and Engineering key laboratory of agricultural genetics and breeding of shanghai Institute of Edible Fungi,shanghai academy of agricultural sciences,shanghai 2.1106,china ~2shanghai Fisheries University,shanghai 200090,P.R.china)

PCR amplification employing a SCAR (Sequence Characterized Amplified Region) primer pair, 135F/R,(amplifying a band of 601 bp) for identifying '135' strains of Lentinula dodes was used to establish the distribution of the specific SCAR marker among 58 protoplast monokaryons and 97 spore monokaryons isolated from a *** 135 *** *** segregated into either A1B1 or A2B2 mating type,and the SCAR marker was detected only in the *** marker was also detected in four delineated mating types (A1B1,A2B2,A1B2,A2B1) of spore *** data testify to the stable inheritance of the specific SCAR marker,and show that the SCAR locus is not genetically linked to either the A or B mating factors.

关键词： Inheritance patterns Lentinula edodes mating type strain-specific sequence characterized amplified region (SCAR) marker

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Analysis of Bacterial Communities Present in Agaricus bisporus Phase II Compost Using Amplified Ribosomal DNA Restriction Analysis (ARDRA)(Ⅰ)——Construction of a Bacterial 16S rDNA Library

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食用菌学报 2008年第4期 16-19页

作者： HE Lihong~(1,2) YU Rongli~1 CHEN Mingjie~(1*) PAN Yingjie~3(~1key laboratory of Applied Mycological Resources and Utilization,Ministry of Agriculture,the People's Republic of china,shanghai research center for Edible Fungi biotechnology and Engineering,shanghai key laboratory of agricultural genetics and breeding,Institute of Edible Fungi,shanghai academy of agricultural sciences,shanghai 2.1106,china ~2Department of Microbiology,College of Life sciences,Nanjing agricultural University,Nanjing,Jiangsu 210095,china ~3shanghai Ocean University,shanghai 200090,china)

An efficient and cost effective method for obtaining good quality DNA from composts containing high levels of organic matter has been *** protocol consisted of sample washing with sodium phosphate and EDTA,cell wall disruption using combined chemical and enzyme treatment prior to extraction,removal of humic acid and other inhibitors in high salt buffer containing polyvinyl polypyrrolidone (PVPP),and precipitation of DNA with polyethylene glycol (PEG-8000).The method was used to extract 16S ribosomal DNA from four samples taken at different stages in the preparation of PhaseⅡcompost used for Agaricus bisporus *** DNA was sufficiently free of contaminants to allow PCR amplification of products derived from bacteria,actinomycetes and *** products derived from PCR amplification of the extracted 16S rDNA using primers F27 and R1498 were used to construct 16S rDNA *** amplification of ten randomly selected positive clones using primer M13 revealed that the 16S rDNA insertion rate was higher than 90%.

关键词： Agaricus bisporus compost phaseⅡ bacterial 16S rDNA library

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Estimating the copy number of transgenes in transformed rice by real-time quantitative PCR

Estimating the copy number of transgenes in transformed rice...

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中国植物生理学会第九次全国会议

作者： Litao Yang, Jiayu Ding, Chengmei Zhang, Junwei Jia, Haibo Weng, Wenxuan Liu, Dabing Zhang (School of life Science and biotechnology, shanghai Jiao Tong University, 800 Dongchuan road, shanghai 200240, P. R. china Department of Biological Science and Technology, Nanjing University, 22 Hankou road, Nanjing 210093, P. R. china key laboratory of agricultural genetics and breeding, agro-biotech research center, shanghai academy of agricultural sciences, 2.01 beidi road, shanghai 2.1106, P.R. china School of Life Science, shanghai University, 99 Shangda road, shanghai 200436, P.R. china)

＜正＞ Abstract: In transgenic plants, the transgene copy number can greatly influence the expression level and genetic stability of the target gene, making estimation of transgene copy number an important part of genetically modified （GM） crop research. Currently, transgene copy numbers are estimated by Southern analysis, which is laborious, time-consuming, requires relatively large amounts of plant materials and may involve hazardous radioisotopes. Here, we report the development of a sensitive, high-throughput real-time PCR technique for estimating transgene copy number in GM rice. This system uses TaqMan quantitative real-time PCR and comparison to a novel rice endogenous reference gene, sucrose phosphate synthase （SPS）, to determine the copy numbers of exogenous β-glucuronidase （GUS） and hygromycin phosphortransferase （HPT） genes in transgenic rice. The GUS and HPT copy numbers in primary rice transformants （T0） were calculated by comparing quantitative PCR results of the GUS and HPT genes with those of the internal standard, SPS. With optimized PCR conditions, we achieved significantly accurate estimates of 1,2,3 and 4 transgene copies in the T0 transformants. Furthermore, our copy number estimations for both the GUS reporter gene and the HPT selective marker gene showed that rearrangements of the T DNA occurred more frequently than is generally believed in transgenic rice.

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Cloning of a Putative Pheromone Receptor Gene Fragment from Lentinula edodes

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食用菌学报 2008年第2期15卷 4-6页

作者： BAO Dapeng ZHANG Meiyan LI Su CHEN Mingjie shanghai research center for Edible Fungi biotechnology and Engineering Key Laboratory of Agricultural Genetics and Breeding of Shanghai Institute of Edible Fungi Shanghai Academy of Agricultural Sciences Shanghai 201106 China

A 508 bp fragment of a putative pheromone receptor gene from Lentinula edodes was amplified by PCR using a pair of degenerate primers designed according to conserved sequences of pheromone receptor genes reported in other *** fragment encodes 117 amino acids and contains three introns,56 bp,51 bp and 50 bp in *** analysis revealed that the deduced amino acid sequence of the *** fragment exhibited 65% identity and 82% similarity with the corresponding segments of pheromone receptors reported in Coprinus cinereus and Schizophyllum commune.

关键词：克隆技术信息素受体基因

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Regulatory diversification of multicopy microRNA family members in plants

Regulatory diversification of multicopy microRNA family memb...

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中国科学院植物分子生物学前沿国际研讨会

作者： Dan-Hua Jiang Chang-Song Yin Ai-Ping Yu Xiao-Fan Zhou Wan-Qi Liang Zheng Yuan Yun Xu Qing-Bo Yu Tie-Qiao Wen Da-Bing Zhang SJTU-SIBS-PSU Joint center for Life sciences School of Life Science and biotechnology shanghai Jiao Tong University 800 Dongchuan road shanghai 200240 P. R. china School of life Science shanghai University 99 Shangda road shanghai 200444 P. R. china College of Life and Environment sciences shanghai Normal University 100 Guilin road shanghai 200234 P. R. china key laboratory of agricultural genetics and breeding agro-biotech research center shanghai academy of agricultural sciences 2.01 beidi road shanghai 2.1106 P. R. china

＜正＞One unique character of miRNA genes in plants is that most of them are predicted to be more than one copy in the genome. About 50 % of identified miRNAs are corresponding to multiple loci in either Arabidopsis or rice genome. Also, 33% of 117 miRNAs from Arabidopsis or 39% of 173 miRNAs from rice were segmental duplications, suggested that genome duplications contributed to the expansion of miRNA families. To analyze the expression pattern of multicopy miRNA family

关键词： gene duplication/microRNA/promoter/RNA polymerase Ⅱ/transcription.

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