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Isolation, Cloning, and Identification of Expressed Sequence Tags from the Backfat Tissue in Duroc and Tongcheng Pigs by mRNA Differential Display

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Agricultural Sciences in China 2006年第2期5卷 141-145页

作者： REN Zhu-qing XIONG Yuan-zhu DENG Chang-yan LEI Ming-gang ZUO Bo LI Feng-e ZHENG Rong XU De-quan Key Laboratory of Swine Breeding and Genetics Ministry of Agriculture/Huazhong Agricultural University Wuhan 430070 P.R.China

mRNA differential display technique was performed to discuss the differential expression of genes in fat tissue between introduced European and Chinese indigenous pigs. Four anchor primers in combination with five arbitrary primers （20 sets in total） were used and nearly 300 bands were observed in polyacrylamide gel, among which 29 differential display bands were obtained. Twelve of 29 cDNA fragments were identified using reverse Northern dot blot, and subsequently cloned and sequenced. Eight of 12 cDNAs had no matches in GenBank and were submitted to GenBank, and the other 4 showed similarity to identified genes from GenBank. Three among 8 novel ESTs were selected to be further identified by semiquantitative rt-pcr. In our experiment, silver staining DDrt-pcr and DIG primer DNA labeling reverse Northern dot blot were used to avoid radioactive pollution. The result showed that the expressions of 5 among 8 novel ESTs were stronger in the backfat of Tongcheng pigs and the others were weaker than that in Duroc pigs. These novel ESTs were prepared for selecting genes related to adipose cells.

关键词： ESTs fat tissue reverse Northern dot blot semi-quantitative rt-pcr mRNA differential display

Isolation, Identification and Tissue Expression Profile Analysis of One Novel Differentially Expressed Sequence Tag in the Longissimus dorsi Muscle from Meishan, Meishan × Large White Hybrid and Large White Pigs

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Agricultural Sciences in China 2004年第11期3卷 856-861页

作者： LIUYong-gang XIONGYuan-zhu DENGChang-yan KeyLaboratoryofSwineGeneticsandBreeding MinistryofAgriculture／HuazhongAgriculturalUniversityWuhan430070P.R.China

In order to detect the molecular mechanism of heterosis in pigs, the mRNA differential display technique was performed to investigate the differences of gene expression in the Longissimus dorsi tissue from Meishan, Meishan × Large White hybrid and Large White pigs with nine 3'-end anchored primers in combination with ten 5'-end arbitrary primers and nearly 3000 reproducible bands were examined. One novel expressed sequence tag (EST4, GenBank accession number: AY553914) that was differentially expressed in Meishan, Meishan× Large White hybrid and Large White pigs was isolated from the Longissimus dorsi muscle tissue and identified through semi-quantitative rt-pcr. BLAST analysis revealed that the 350 bp long EST (EST4) was not homologous to any of the known porcine genes. Tissue expression profile analyses showed that the EST4 was expressed in most of *** Yong-gang, Ph D candidate

关键词： mRNA differential display semi-quantitative rt-pcr Tissue expression profile analysis

Isolation, Identification of Differentially Expressed Sequence Tags in the Backfat Tissue from Meishan, Large White and MeishanLarge White Cross Pigs

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Agricultural Sciences in China 2005年第1期4卷 54-58页

作者： LIUYong-gang XIONGYuan-zhu DENGChang-yan KeyLaboratoryofSwineGeneticsandBreeding／MinistryofAgriculture HuazhongAgriculturalUniversityWuhan430070P.R.China

In order to detect the molecular mechanism of heterosis in pigs, the mRNA differential display technique was performedto investigate the differences of gene expression in the backfat tissue from Meishan, Large White and MeishanLargeWhite cross pigs. Nine 3'-end anchored primers in combination with ten 5'-end arbitrary primers were used to perform thedifferential display pcr and nearly 3 000 reproducible bands were examined. Fifteen expressed sequence tags that weredifferentially expressed were isolated and then identified through semi-quantitative rt-pcr. BLAST analysis revealedthat the fifteen expressed sequence tags (ESTs) were not homologous to any of the known porcine genes or ESTs. Thesenovel ESTs were then submitted to GenBank.

关键词： Pig mRNA differential display semi-quantitative rt-pcr Expressed sequence tag

Distribution and Localization of Gonadotropin-inhibitory Hormone( GnIH ) in the Central Nervous System of Male Luchuan Piglets

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Animal Husbandry and Feed Science 2016年第2期8卷 63-67,78页

作者： Huo Konglin Hu Hao Li Jianhua Luo Bo Wang Xiaoye Li Xun College of Animal Science and Technology Guangxi University

Immunohistochemistry （IHC） and semi-quantitative rt-pcr were used to detect the distribution of GnIH in the central nervous system of 30-day-old male Luchuan piglets, which laid a theoretical basis for in-depth research on physiological functions of Gn[H from the perspective of morphology and molecular lev- el. IHC results showed that in central nervous system, GnIH immunoreactive neurons distributed in cerebral cortex, cerebellum, hippocampus, olfactory bulb, ports, midbrain, medulla oblongata and spinal cord of Luchuan piglets, but GnIH-immunoreactive （GnIH-ir） cell bodies employed diversified morphologies with va- rying sizes. The results of semi-quantitative rt-pcr showed that the expression of GnIH mRNA in medulla obhingata was the highest, followed by midbrain, and the expression of GnIH mRNA in olfactory bulb was the lowest, while those in other tissues were medium. The results of IHC and semi-quantitative rt-pcr were coincident, which further confirmed wide distribution of GnIH in the central nervous system of male Luchuan piglets.

关键词： Luchuan piglet GnIH Immunohistochemistry semi-quantitative rt-pcr

Cloning of Ds MAPK Gene and Construction of Antisense Expression Vector

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Agricultural Biotechnology 2018年第4期7卷 8-11,24页

作者： Jinrong YUE Yuting CONG Xiangnan GAO Zhenyu XING Wenjing YUE Xiaojie CHAI Dalian Ocean University/Key Laboratory of Maricuhure&Stock Enhancement in North China's Sea Ministry of Agricuhure/Key Laboratory of Marine Bio-Resources Restoration and Habitat Reparation in Liaoning ProvinceDalian 116023China

In order to investigate the role of MAPK gene in adaptation of Dunaliella salina to hypersaline environment, the Ds MAPK gene of D. salina was amplified by pcr. After inverted insertion of the open reading flame ofDs MAPK gene into downstream sequence of 35S promoter of plant expression vector, an antisense expression vector of Ds MAPK gene was successfully constructed and introduced into D. salina cells by LiAc/PEG-mediated method. The expression of Ds MAPK gene in transgenic D. salina was analyzed by semi-quantitative rt-pcr method. The results showed that the expression of Ds MAPK gene in D. salina was significantly inhibited at the transcriptional level. The study laid the foundation for further identification of the function of Ds MAPK gene.

关键词： Dunaliella salina Ds MAPK Antisense expression vector semi-quantitative rt-pcr

Effects of cold stress on hypothalamic corticotrophin-releasing and thyrotropin-releasing hormone messenger RNA levels in chickens

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Journal of Northeast Agricultural University(English Edition) 2007年第4期14卷 327-332页

作者： WANG Jintao XU Shiwen College of Veterinary Medicine Northeast Agricultural University Harbin 150030 China Animal Science and Veterinary Medicine Research Institute Heilongjiang Academy of Nongken Sciences Harbin 150038 China

There is increasing evidence that stress can activate the hypothalamic-pituitary-adrenal-axis and hypothalamic-pituitary- thyroid-axis, and further affect the synthesis and secretion of corticotrophin-releasing hormone （CRH） and thyrotropin-releasing hormone （TRH）. To evaluate the effect of cold stress on the hypothalamic CRH and TRH messenger RNA （mRNA） levels in Yisha chickens, male Yisha chickens were subjected to acute （1, 6, 12 h） and chronic （5, 10, 20 d） cold stress （12±1）℃. Hypothalami were collected for assessment of mRNA levels by semi-quantitative rt-pcr. Acute stress resulted in a significant decrease of CRH mRNA levels at 6 and 12 h, and a significant increase of TRH mRNA levels at every stress time point. Chronic cold stress resulted in a significant increase of CRH mRNA levels and a significant decrease of TRH mRNA levels compared with the control group at every stress time point. The results suggest that the two genes differently respond to cold stress at the mRNA levels. And the different degrees of cold stress will produce different effects on the identical gene.

关键词： chicken cold stress semi-quantitative rt-pcr hypothalamus CRH mRNA TRH mRNA

Cloning,Expression and Bioinformatic Analysis of Dorsal Gene in Delia antique

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Plant Diseases and Pests 2012年第2期3卷 28-32页

作者： Huimin WEI Yujuan ZHANG Bin CHEN Shuang REN Institute of Entomology and Molecular Biology Chongqing Engineering Research Center of Bioactive Substances Chongqing Key Laboratory of Animal Biology Chongqing Normal University

[ Objective ] The aim was to explore the relationship between the Dorsal and diapause development of Delia araiqua. [ Method ] The full-length cDNA of Dorsal in D. antiqua was cloned through RACE. The similarity among deduced amino acid sequence of Dorsal cDNA and the Dorsals of other 14 insect species were compared, and the phylogenetic analysis of these Dorsals was conducted. The expression level of Dorsal gene in winter-, summer- and non-diapausing pupae was analyzed. [ Result] The full-length of Dorsal cDNA sequence, 2 412 bp, was obtained with ORF 1974 bp, which coded 657 amino acids with predicted Mw 72.9 kDa and PI 8.5. The result of similarity comparison indicated that the DaDorsal was most related to those of Drosophlia melanogaster and Drosophlia pseudoobscura. The result of semi-quantitative rt-pcr analysis showed the expression level of Dorsal gene increased in characteristic duration of winter-, summer- and non- diapansing pupae, especially at the late diapanse, which might imply its relationship to D. antiqua diapause and development. [ Conclusion] The study lays the foundation for further study on gene function of Dorsal in insect diapause and development.

关键词： Dorsal Delia ant/qua RACE Sequence analysis Phylogenetic analysis semi-quantitative rt-pcr

Gene Expressing Difference in Sclerotial Formation of Morchella conica

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Gene Expressing Difference in Sclerotial Formation of Morche...

The differential expressed genes of banana induced by Fusarium oxysporum *** infection

2017第二届全国羊肚菌大会暨湖北省食用菌协会第八次会员代表大会

作者： Li-Jiao Chen Hong-Mei Chai Wei-Min Chen Xing-Qi Huang Yong-Chang Zhao

The difference of gene expression betweensclerotia-producing and non-sclerotia producing singlespore isolates from Morchella conica were preliminaryanalyzed by mRNA differential display reverse transcrip-tionpolymerase chain reaction(rt-pcr) technique and 67 differential gene fragments were ***-eight oftheir second pcr products were cloned and *** special differential gene fragments related tosclerotial formation were validated by semiquantitative *** gene fragments had certain homologies with lipoprotein,cyclin-dependent kinase C-3,glycerophosphoryl diester phosphodiesterase,Rho GDPdissociation inhibitor,gamma-aminobutvrate permease,OmpA family protein,Transcript antisense to ribosomal RNA protein,sodiumcalcium exchange protein and keratinassociated proteins 5,*** addition,the putative protein of some DNA fragments had higher similarity with hypothetical proteincoding gene in NCBI database,as well as some were only putative gene *** these fragments were speculated to be the functional gene associated with sclerotial formation in morel.

关键词： mRNA differential display Morchella conica Sclerotia semi-quantitative rt-pcr

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The differential expressed genes of banana induced by Fusari...