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Production of human lysozyme-transgenic cloned porcine embryos by somatic nuclear transfer

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Progress in Natural science:Materials International 2009年第6期19卷 699-704页

作者： Qiuyan Lia,b, Hengxi Wei a, Ying Guoa, Yan Lib, Rui Zhao b, Yufang Maa, Zhengquan Yu a, Bo Tang b, Lei Zhangb, Yunping Dai a,b, Ning Li a, a State Key Laboratory for Agrobiotechnology and department of animal Physiology, college of Biological science, china agricultural university, No. 2 Yuan-Ming-Yuan West Road, beijing 100094, china b Ji Pulin Biotech Ltd, beijing 100094, china State Key Laboratory for Agrobiotechnology and department of animal Physiology College of Biological Science China Agricultural University No. 2 Yuan-Ming-Yuan West Road Beijing 100094 China Ji Pulin Biotech Ltd Beijing 100094 China

Due to their physiology and organ size, pigs have significant potential as human disease models and as organ transplantation donors. Genetic modification of pigs could provide benefits for both agriculture and human medicine. In this study, five fetal pig fibroblast cell lines from two species (Wuzhishan and Landrace pigs) were transfected using double-marked human lysozyme (HLY) plasmids (pBC1-HLY-GFP-NEO) by a liposome-mediated method. The ratio of green fluorescent protein (GFP)-expressing cells was >95% in sw7, sw8, slw3 and slw6 cell lines, but only 49.3% in slw9 cells. Cells from the four highly transgenic lines were used as nuclear donors to construct embryos, which were then cultured after fusion and activation by electric stimulation. The rate of cleavage was 76.7%, 48 h after activation. After 7 days, 18.5% of cleaved eggs had developed to the blastocyst stage and 93.3% of blastocysts were GFP-positive. These results indicate that transgenic fetal pig flbroblast cell lines could be obtained by a liposome-mediated method, though the transfection efficiency varied between cell lines. Reconstructed embryos derived from transgenic cells could successfully develop into blastocysts, most of which were GFP-positive.

关键词： EGFP Porcine Nuclear transfer Fetal fibroblast Human lysozyme (HLY)

Possible Effect of 30K Proteins in Embryonic Development of Silkworm Bombyx mori

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Acta Biochimica et Biophysica Sinica 2005年第5期37卷 355-361页

作者： Bo-Xiong ZHONG~1 Jian-Ke LI~(1.2) Jian-Rong LIN~3 Jian-She LIANG~4 Song-Kun SU~1 Hai-Sheng XU~1 Hai-Yan YAN~1 Ping-Bo ZHANG~5 Hiroshi FUJII~5 1 Laboratory of Molecular Biology of Silkworm and Honeybee, department of Bioresource science, college of animal science,Zhejiang university, Hangzhau 310029, china 2 Institute of Apicultural Research, Chinese Academy of agricultural science, beijing 100093, china 3 department of Sericulture, South china agricultural university.Guangzhou 510642, china 4 Biochemistry Co. Ltd. of Zhejiang university, Hangzhou 310029. china 5 Faculty of Agriculture,Kyushu university, Fukuoka 812-81. Japan Laboratory of Molecular Biology of Silkworm and Honeybee Department of Bioresource Science College of Animal Science Zhejiang UniversityHangzhou 310029 China Institute of Agicultural Research Chinese Academy of Agricultural ScienceBeijing 100093 China department of Sericulture South China Agricultural UniversityGuangzhou 510642 China Biochemistry Co. Ltd. of Zhejiang universityHangzhou 310029 China Faculty of Agriculture Kyushu UniversityFukuoka 812-81 Japan

The silkworm Bombyx mori possesses a 30K protein family of 3×10～4 Da,the biologicalfunctions of which have not been fully *** relationship between the 30K protein family and theembryonic development of temperature sensitive sex-linked mutant strain of silkworm was investigated bytwo dimensional polyacrylamide gel electrophoresis(2D-PAGE)and Matrix assisted laser desorption ionization-time of flight mass spectrometry(MALDI-TOF MS).The results show that protein spots 1-5 of the 30Kprotein family,mainly existing in normal strain,are possibly related to embryonic *** earlyconsumption of a 30K protein named 6G1-30K-1 and the accumulation of 30K proteins named 6G1-30K-3and 6G1-30K-4 are likely caused by the destruction of physiological balance in normal embryonic development,which may lead to lower hatchability of the temperature sensitive *** results suggest that reasonablemetabolism of 30K proteins is a prerequisite for the embryo's normal development.

关键词： silkworm Bombyx mori 30K protein embryonic development 2D-PAGE MALDI-TOF MS

Soluble expression and characterization of a GFP-fused pea actin isoform (PEAc1)

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Cell Research 2004年第5期14卷 407-414页

作者： Ai Xiao LIU*, Shao Bin ZHANG*, Xiao Jing XU, Dong Tao REN, Guo Qin LIU** State Key Laboratory of Plant Physiology and Biochemistry, college of Biological science, china agricultural university, beijing 100094, china. State Key Laboratory of Plant Physiology and Biochemistry College of Biological Science China Agricultural University Beijing 100094 China

A pea actin isoform PEAc1 with green fluorescent protein (GFP) fusion to its C-terminus and His-tag to its N- terminus, was expressed in prokaryotic cells in soluble form, and highly purified with Ni-Chelating SepharoseTM Fast Flow column. The purified fusion protein (PEAc1-GFP) efficiently inhibited DNase I activities before polymerization, and activated the myosin Mg-ATPase activities after polymerization. The PEAc1-GFP also polymerized into green fluorescent filamentous structures with a critical concentration of 0.75 μM. These filamentous structures were labeled by TRITC-phalloidin, a specific agent for staining actin microfilaments, and identified as having 9 nm diameters by negative staining. These results indicated that PEAc1 preserved the essential characteristics of actin even with His-tag and GFP fusion, suggesting a promising potential to use GFP fusion protein in obtainning soluble plant actin isoform to analyze its physical and biochemical properties in vitro. The PEAc1-GFP was also expressed in tobacco BY2 cells, which offers a new pathway for further studying its distribution and function in vivo.

关键词： actin isoform polymerization DNase I inhibition myosin Mg-ATPase activation expression.

GENETIC CHANGES AFFECTING POULTRY PRODUCTION IN china

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GENETIC CHANGES AFFECTING POULTRY PRODUCTION IN CHINA

Identification and mapping of quantitative trait loci controlling cold-tolerance of Chinese common wild rice (O. rufipogon Griff.) at booting to flowering stages

面向21世纪的动物科学与动物医学国际研讨会

作者： Changxin Wucollege of animal science and technology china agricultural university beijing 100094 china.

＜正＞Genetic changes in poultry production in china are due to theory, method and resources. Theory of quantitative genetics, breeding methods and technology, a diversity of poultry genetic resources made its possible to get genetic improvement during the past 20 years. It is also important to establish a breeding system for poultry production. A great number of poultry-men get benefits from commercial stock with high genetic merits. State breeding farms keep seed stock with pedigree, and select birds according to the breeding program.

关键词： genetic changes poultry production china

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Chinese science Bulletin 2003年第19期48卷 2068-2071页

作者： LIU Fengxia, SUN Chuanqing, TAN Lubin, FU Yongcai, LI Dejun & WANG Xiangkun department of Plant genetics and breeding, china agricultural university, National Key Laboratory of Agrobiology, Key Laboratory of Crop Genetic Improvement and Genome of the Ministry of Agriculture, beijing 100094, china department of Plant genetics and breeding China Agricultural University Beijing China National Key Laboratory of Agrobiology Key Laboratory of Crop Genetic Improvement and Genome of the Ministry of Agriculture Beijing China

An advanced backcross population of rice was used to identify the quantitative trait locus (QTL) controlling the cold-tolerance at booting to flowering stages. The recipient, Guichao 2 (GC2), was a commercial Indica rice; the donor Dongxiang common wild rice, was an accession of common wild rice (DXCWR, Oryza rufipogon Griff.). Three QTLs for cold-tolerance were detected on chromosomes 1, 6 and 11. Two of them coming from DXCWR could enhance the cold-tolerance of the backcross progenies. Moreover, one sterility QTL that could reduce the seed set rate of the backcross progenies by 78% was mapped on chromosome 5.

关键词：耐寒性水稻开花期播种设备回交杂种 QTL分析数量特点轨迹

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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

A Comparative Analysis of Molecular Diversity and Population Structure of Hemarthria compressa（whipgrass）in South china Using ISSR and SRAP markers

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A Comparative Analysis of Molecular Diversity and Population...

Mapping QTLs on BTA6 affecting milk production traits in a Chinese Holstein population

The 3rd Korea-china-Japan Joint Symposium on Grassland Agriculture and Livestock Production

作者： Linkai Huang~1,Xinquan Zhang~(1*),Surong Shuai~2,and Fang Li~1 1 department of Grassland science,college of animal science and technology,Sichuan agricultural university,Yaan,625014,china 2.department of animal genetics and breeding,college of animal science and technology,Sichuan agricultural university,Yaan,625014,china

＜正＞Genetic variation within and among 12 populations of whipgrass in south china were investigated using inter-simple sequence repeat（ISSR） and sequence-related amplified polymorphism（SRAP）.High genetic diversity was found in whipgrass by two molecular makers（PPB=86.21%,I=0.357 based on ISSR;PPB=82.21%,I=0.352 based on SRAP）.However,there was relatively low genetic diversity at population levels.A high degree of genetic differentiation among populations was detected based on different measures and different molecular *** also found that SRAP markers were more efficient than ISSR markers in this *** on these findings,sampling strategy was proposed for successfully utilizing the genetic resource of this species.

关键词： Hemarthria compressa ISSR SRAP Genetic diversity Population structure

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science Bulletin 2005年第16期 67-72页

作者： CHEN Huiyong1, ZHANG Qin2, WANG Chunkao1, SHU Juan1, MEI Gui1, YIN Cengceng1, HU Fang1, XU Jingjing1, GONG Weijia1, LI Hejun1 & QIU Xiaotian1 1. Key Laboratory of animal genetics and breeding of the Ministry of Agriculture, college of animal science and technology, china Agri-cultural university, beijing 100094, china 2. college of animal science and technology, china agricultural Uni-versity, beijing 100094, china

A Chinese Holstein population with daughter design was analyzed using 14 microsatellites covering a map distance of 55.7 cM on chromosome 6 to fine map QTL for five milk production traits. 26 paternal half-sib families with 2356 daughters were involved. Two different approaches, linear regression approach and variance component ap-proach, were employed, with a one-QTL model and two-QTL model fitted. With a one-QTL model, the linear regression approach revealed a QTL near BMS470 with effects on milk yield, fat yield, protein yield, and fat percentage, and another QTL near BMS2460 for protein percentage. The variance component approach confirmed the results of linear regres-sion approach for the three yield traits, with the exception that the QTL for fat yield was mapped to a different position near BMS1242. The 95% confidence intervals resulted from linear regression, obtained by bootstrapping, were generally large, ranging from 31 to 53 cM, whereas the variance com-ponent approach revealed very small confidence intervals, calculated by LOD drop-off method, for the three yield traits, only 4―5 cM. With a two-QTL model, both approaches pro-vided strong evidence for the existence of two QTLs for the three yield traits. Along with the QTLs identified in one-QTL model analyses, the linear regression approach revealed a second QTL near BP7 with effects on all the three yield traits, whereas the variance component approach located the sec-ond QTL near ILSS035, BMS470, and BP7 for the three traits, respectively.

关键词： Chinese Holstein cattle daughter design milk produc-tion trait linear regression approach variance component approach.

Cloning and expression profiles of 15 genes encoding WRKY transcription factor in wheat (Triticum aestivem L.)

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Progress in Natural science:Materials International 2008年第6期18卷 697-705页

作者： Hualing Wu,Zhongfu Ni,Yingyin Yao,Ganggang Guo,Qixin Sun* department of Plant genetics&breeding and State Key Laboratory for Agrobiotechnology,Key Laboratory of Crop Heterosis and Utilization,Ministry of Education,Key Laboratory of Crop Genomics and Genetic Improvement,Ministry of Agriculture/beijing Key Laboratory of Crop Genetic Improvement,china agricultural university,beijing 100094,china department of Plant genetics & breeding and State Key Laboratory for Agrobiotechnology Key Laboratory of Crop Heterosis and Utilization Ministry of Education Key Laboratory of Crop Genomics and Genetic Improvement Ministry of Agriculture/Beijing Key Laboratory of Crop Genetic Improvement China Agricultural University Beijing 100094 China

WRKY proteins are involved in various physiological processes,including biotic and abiotic stress responses,hormone responses and ***,no systematic identification,expression and function analysis of WRKY genes in wheat were *** this study,we isolated 15 wheat cDNAs with complete open reading frame(ORF)encoding putative WRKY proteins using in silico *** analysis indicated that the 15 wheat WRKY genes belonged to three major WRKY *** analysis revealed that most genes expressed drastically in leaf,except TaWRKY10 which expressed in crown *** genes were strongly up-reg-ulated with the senescence of *** genes were responsive to low temperature,high temperature,NaCl or PEG ***-over,dierential expression patterns were also observed between wheat hybrid and its parents,and some genes were more responsive to PEG treatment in the *** results demonstrated that wheat WRKY genes are involved in leaf senescing and abiotic *** the changed expression of these WRKY genes in hybrid might contribute to the heterosis by improving the stress tolerance in hybrids.

关键词： Wheat WRKY transcription factor Gene cloning Gene expression Heterosis