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Progress in Natural Science:Materials International 2006年第4期16卷 366-372页

作者： XIE Baoen and CHEN Sanfeng State key laboratory for agrobiotechnology, College of Biological Sciences,{3., beijing 100094, china State key laboratory for agrobiotechnology College of Biological Sciences China Agricultural University Beijing 100094 China

To bring more information about synthesis of indole-3.acetic acid (IAA) from Azospirillum brasilense, a Tn5-insertion library of A. brasilense Yu62 was constructed and subjected to screening for IAA producing mutants. Two mutants with decreased IAA levels, named as A3.and A24, were isolated. The sequence analysis of loci tagged showed that the Tn5-1063. was located in the atrA gene encoding GntR family transcriptional regulator and trpE gene encoding component I of anthranilate synthase respectively. At the same time. atrB encoding phosphotransferase and atrC encoding aminotransferase were cloned downstream the atrA gene and atrA, atrB and atrC were clustered in an operon. Mutagenesis and complementation studies showed that atrA and atrC were involved in IAA synthesis. IAA levels of trpE mutant and wild-type strain could be improved by adding anthranilate into the medium.

关键词： indole-3.acetic acid (IAA) IAA mutants Azospirillum brasilense atrABC cluster trpE Tn5 transposon.

Genetic analysis on 3.- terminal flanking region of uncoupling protein 3.in different pig breeds

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Chinese Science Bulletin 2002年第18期47卷 1541-1543页

作者： FANG Meiying, ZHAO Xingbo, LI Ning & WU ChangxinCollege of Animal Science and Technology, china agricultural university, beijing 100094, china State {3. laboratory for agrobiotechnology, china agricultural university, beijing 100094, china College of Animal Science and Technology China Agricultural University Beijing China State key laboratory for agrobiotechnology China Agricultural University Beijing China

The 3.-terminal flanking region of porcine uncoupling protein 3.(UCP3. was cloned, the sequence data revealed 15 nucleotide substitutions among Landrace and three Chinese native pig breeds named Neijiang, Minpig and Erhualian. The continuous 9 polymorphic sites were checked by PCR-RFLP, the results indicated that Erhualian had extraordinary gene frequency, presented most significant difference by x2 test compared with Landrace, Largewhite, Neijiang and Minpig respectively, significant level compared with Meishan; and Meishan also had significant difference compared with Landrace and Minpig respectively. These results can be concluded that Taihu pigs have special genetic characteristics among pig breeds.

关键词： pig uncoupling protein gene clone PCR-RFLP.

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Cloning and sequencing of nifBHDKENX genes of Paenibacillus massiliensis T7 and its nif promoter analysis

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Science china(Life Sciences) 2006年第2期49卷 115-122页

作者： ZHAO Hongxin1,2,3. XIE Baoen2 & CHEN Sanfeng1,2 1. State key laboratory for agrobiotechnology and Biological School,{3., beijing 100094, china 2. key laboratory for Agro-Microbial Resource and Application of Agriculture Ministry,{3., beijing 100094, china 3. Chemical and Biological School, ShenYang Normal university, Shenyang 11003., china 1. State key laboratory for agrobiotechnology and Biological School China Agricultural University Beijing 100094 China 2. key laboratory for Agro-Microbial Resource and Application of Agriculture Ministry China Agricultural University Beijing 100094 China 3. Chemical and Biological School ShenYang Normal University Shenyang 110034 China

A 3.4 bp of nifH fragment was PCR amplified from Paenibacillus massiliensis T7 using the universal degenerate primers. The PCR-amplified nifH fragment was labeled with DIG; then used as a probe in Southern blot analysis. Southern blot result showed that there were two positive signals, indicating that there might be two copies of nifH in P. massiliensis T7. A total of 10254 bp DNA sequence containing purD; nifBHDKENX was obtained by five rounds of inverse-PCR amplification. The predicted proteins of nifBHDKENX had high homology with those from other nitrogen-fixing bacteria. Only one putative σ[54]-dependent promoter sequence was detected upstream of the nifB gene; nifBHDKENX were likely to be organized in one operon. Assays of β-galactosidase activity of P. massiliensis T7PB carrying a nifB-lacZ fusion under different concentrations of NH4 [+]; O2 showed that the expression of nifB-lacZ was strongly inhibited by O2.

关键词： Paenibacillus massiliensis T7, nifBHDKENX, nifB-lacZ, inverse-PCR.

维普期刊数据库

Aberrant histone H4 acetylation in dead somatic cell-cloned calves

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Progress in Natural Science:Materials International 2008年第7期18卷 813-818页

作者： Lei Zhang a,b,*, Shaohua Wang a, Qiang Li a, Xiangdong Ding c, Yunping Dai a, Ning Li a,* a State key laboratory for agrobiotechnology,{3., beijing 100094, china b beijing Gene & Protein Biotechnology Co. LTD, beijing 100094, china c College of Animal Science and Technology,{3., beijing 100094, china State key laboratory for agrobiotechnology China Agricultural University Beijing 100094 China beijing Gene & Protein Biotechnology Co. LTD Beijing 100094 China College of Animal Science and Technology China Agricultural University Beijing 100094 China

In somatic cell-cloned animals, inefficient epigenetic reprogramming can result in an inappropriate gene expression and histone H4 acetylation is one of the key epigenetic modifications regulating gene expression. In this study, we investigated the levels of histone H4 acetylation of 11 development-related genes and expression levels of 19 genes in lungs of three normal control calves and nine aber- rant somatic cell-cloned calves. The results showed that nine studied genes had decreased acetylation levels in aberrant clones (p 0.05). Whereas 13.genes had significantly decreased expression (p 0.05), and only one gene had higher expression level in clones (p < 0.05). Furthermore, FGFR, GHR, HGFR and IGF1 genes showed lowered levels of both histone H4 acetylation and expression in aberrant clones than in controls, and the level of histone H4 acetylation was even more lowered in aberrant clones than those in controls. It was suggested that the lower levels of histone H4 acetylation in aberrant clones caused by the previous memory of cell differentiation might not support enough chromatin reprogramming, thus affecting appropriate gene expressions, and growth and development of the cloned calves. To our knowledge, this is the first study on how histone H4 acetylation affects gene expression in organs of somatic cell-cloned calves.

关键词： Somatic cell-cloned calves Histone H4 acetylation Gene expression Lungs

Cloning and characterization of an up-regulated GA 20-oxidase gene in hybrid maize

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Progress in Natural Science:Materials International 2009年第2期19卷 161-166页

作者： Jinkun Du,Yingyin Yao,Zhongfu Ni,Qixin Sun Department of Plant Genetics & Breeding and State key laboratory for agrobiotechnology,key laboratory of Crop Genomics and Genetic Improvement,Ministry of Agriculture/beijing key laboratory of Crop Genetic Improvement,china agricultural university,beijing 100094,china [a]Department of Plant Genetics & Breeding and State key laboratory for agrobiotechnology Key Laboratory of Crop Genomics and Genetic Improvement Ministry of Agriculture/Beijing Key Laboratory of Crop Genetic Improvement China Agricultural University Beijing 100094 China

Previous studies revealed that GA content and metabolism are positively correlated with a faster shoot growth rate of hybrid,and recently,genes participating in both GA biosynthesis and GA response pathways were also found to be differentially expressed between wheat hybrid and its parental inbreds. In this study,an up-regulated GA 20-oxidase gene in a maize hybrid,designated ZmGA20,was cloned. ZmGA20 contains an open reading frame (ORF) encoding 3.1 amino acid residues. BLASTX searches in GenBank revealed that the ZmGA20 is homologous to the sequences of GA20ox proteins from different species,and analysis also indicated that ZmGA20 had typical features of GA 20-oxidase proteins,including a "LPWKET" sequence. Semi-quantitative RT-PCR analysis showed that ZmGA20 was expressed in different tissues and/or organs. The expression level of ZmGA20 in the hybrid was higher than that in two parents (in roots,leaves,stems and embryo,and ears). The abundance of ZmGA20 transcript was equal to that of the highly expressed parents,which provided molecular evidence for the observed GA content heterosis in maize hybrids.

关键词： GA 20-oxidase Gene expression Maize Heterosis

Signal Peptide of Potato PinII Enhances the Expression of Cry1Ac in Transgenic Tobacco

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Acta Biochimica et Biophysica Sinica 2004年第8期36卷 553-558页

作者： Yun-Jun LIU, Yuan YUAN, Jun ZHENG, Ya-Zhong TAO, Zhi-Gang DONG,Jian-Hua WANG, and Guo-Ying WANGState key laboratory for agrobiotechnology and National Center for Maize Improvement,{3., beijing 100094, china State key laboratory for agrobiotechnology and National Center for Maize Improvement China Agricultural University

The modified Cry1Ac was expressed in transgenic tobacco plants. To allow secretion of the Cry1Ac protein into the intercellular space, the signal peptide sequence of potato proteinase inhibitor II (pinII) was N-terminally fused to the Cry1Ac encoding region. Expression of Cry1Ac in transgenic tobacco plants was assayed with ELISA. The results showed that pinII signal peptide sequence enhanced the expression of Cry1Ac protein and led to the secretion of the Cry1Ac protein in transgenic tobacco plants. GFP gene was also fused to the signal peptide sequence and transformed to tobacco. The results of fluorescent detection showed that GFP had localized in the apoplast of transgenic plants.

关键词： Cry1Ac signal peptide GFP transgenic tobacco

Protein phosphorylation is involved in the water stress-induced ABA accumulation in the roots of Malus hupehensis Rehd

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Chinese Science Bulletin 2001年第10期46卷 855-858页

作者： ZHANG Dapeng, YANG Hongqiang, JIA Wensuo & HUANG Conglin1. laboratory of Molecular Developmental Biology of Fruit Trees, china agricultural university, beijing 100094,{3. 2. The key laboratory of Plant Physiology and Biochemistry of agricultural Ministry, china agricultural university, beijing 100094,{3. laboratory of Molecular Developmental Biology of Fruit Trees China Agricultural University Beijing China The key laboratory of Plant Physiology and Biochemistry of agricultural Ministry China Agricultural University Beijing China

Water stress-induced ABA accumulation plays a key role in the root to shoot communication and/or the cell to cell signaling under the soil stresses. The signaling of the water stress itself that leads to the accumulation, however, is less known. In this study, we subjected the roots of Malus hupehensis seedlings to water stress treatment and investigated the ABA accumulation in relation to protein phos-phorylation. Our results showed that ABA accumulation could be substantially triggered in 40 min and reached 4 folds in 100 min after treatment with 3.% PEG 6000 (polyethylene glycol). The water stress treatment also led to a substantial enhancement of total kinase activity, assessed with histone-111 as substrate, in 15 min and a maximum enhancement in 3. min before it declined to initial level. The Ca2+-dependent kinase activity showed a similar, if not more sensitive, trend. When the roots were fed with labeled 3.P-ATP, water stress enhanced the labeling of proteins, which showed a maximum labeling at

关键词： ABA (abscisic acid) water stress protein kinases Malus hupehensis Rehd.

维普期刊数据库

Genomic and genic sequence variation in synthetic hexaploid wheat (AABBDD) as compared to their parental species

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Progress in Natural Science:Materials International 2008年第5期18卷 533-538页

作者： Lihong Nie, Zongfu Han, Lahu Lu, Yingyin Yao, Qixin Sun, Zhongfu Ni key laboratory of Crop Heterosis and Utilization (MOE) and State key laboratory for agrobiotechnology, key laboratory of Crop Genomics and Genetic Improvement (MOA), beijing key laboratory of Crop Genetic Improvement, Department of Plant Genetics & Breeding, china agricultural university, beijing 100094, china key laboratory of Crop Heterosis and Utilization (MOE) and State key laboratory for agrobiotechnology Key Laboratory of Crop Genomics and Genetic Improvement (MOA) Beijing Key Laboratory of Crop Genetic Improvement Department of Plant Genetics & Breeding China Agricultural University Beijing 100094 China

In order to understand the genomic changes during the evolution of hexaploid wheat, two sets of synthetic hexaploid wheat from hybridization between maternal tetraploid wheat (AABB) and paternal diploid goat grass (DD) were used for DNA-AFLP and single strand conformation polymorphism (SSCP) analysis to determine the genomic and genic variation in the synthetic hexaploid wheat. Results indicated that more DNA sequences from paternal diploid species were eliminated in the synthetic hexaploid wheat than from maternal tetraploid wheat, suggesting that genome from parental species of lower ploidity tends to be eliminated preferentially. However, sequence variation detected by SSCP procedure was much lower than those detected by DNA-AFLP, which indicated that much less variation in the genic regions occurred in the synthetic hexaploid wheat, and sequence variations detected by DNA-AFLP could be derived mostly from non-coding regions and repetitive sequences. Our results also indicated that sequence variation in 4 genes can be detected in hybrid F1, which suggested that this type of sequence variation could be resulted from distant hybridization. It was interesting to note that 3.out of the 4 genes were mapped and clustered on the long arm of chromosome 2D, which indicated that variation in genic sequences in synthetic hexaploid wheat might not be a randomized process.

关键词： Synthetic hexaploid wheat Polyploidy Genome Sequence variation

Aberrant DNA methylation patterns in cultured mouse embryos

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Progress in Natural Science:Materials International 2005年第12期15卷 1079-1083页

作者： HOU Jian,CUI Xiuhong,LEI Tinghua,LIU Lei,AN Xiaorong and CHEN Yongfu State key laboratory for agrobiotechnology and Department of Animal Physiology,College of Biological Science,{3. agricultural university,beijing 100094,{3. State key laboratory for agrobiotechnology and Department of Animal Physiology College of Biological Science China Agricultural University Beijing 100094 China

Mouse early embryos undergo genome-wide demethylation and remethylation events during pre-implantation *** methylation reprogramming is thought to be associated with development *** immunofluorescence staining with an antibody against 5-methylcytosine (MeC),we examined the genome methylation patterns of mouse embryos cultured in *** results did not show the difference in staining patterns between development-blocked two-cell embryos that cultured in vitro and the two-cell embryos that were freshly collected from the donor *** in vitro-arrested morulae displayed a strong positive staining when compared to the morulae freshly collected from the donor *** the blastocyst stage,although most embryos showed the expected methylation patterns,with highly stained inner cell mass (ICM) and weekly stained trophectoderm (TE),a proportion of embryos were dimly stained in both ICM and *** results indicated that the methylation profile of the embryos could be changed by culturing in vitro when the embryos were in the transition from morulae to blastocyst.

关键词： methylation mouse embryo 5-methylcytosine antibody

A five-fold pig bacterial artificial chromosome library:a resource for positional cloning and physical mapping

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Progress in Natural Science:Materials International 2006年第08期16卷 889-892页

作者： LIU Wei, ZHANG Ying, LIU Zhaoliang, GUO Li, WANG Xiaobo, FEI Jing, FENG Jidong, ZHAO Rui, HU Xiaoxiang and LI Ning (The State key laboratory for Agro-Biotechnology, College of Biology, china agricultural university,{3., china College of Animal Science and Technology, china agricultural university,{3., china) [a]The State key laboratory for Agno-Biotechnology College of Biology China Agricultural University Beijing China [b]College of Animal Science and Technology China Agricultural University Beijing China

A pig BAC library was constructed with genomic DNA from a male Erhualian pig. After partial digestion with HindⅢor BamHⅠthe fragments obtained were cloned into the pBeloBAC11 vector. The library consists of 1843.0 clones which stored in 480 pieces 3.4-well plates (20 plates per superpool). A two-step 4-dimension PCR screening system was established to screen the positive clones. An average insert size of 128 kb was estimated from 105 randomly isolated clones, which indicates that the library is more than five times of genomic coverage. For the demonstration of the probability to pick out any unique genes or DNA markers from the library, 10 single-copy genes were screened out and the positive clones were yielded between 1 and 8 with an average of 3.6. Positive superpools were obtained for 3. microsatellite markers selected from different regions of pig genome. The number of positive superpools for each marker varies from 1 to 9 with an average of 4.78. This BAC library provides an additional resource for pig physical mapping and gene identifica-

关键词： pig BAC library PCR screening system.