Background Photosystem II(PSII)constitutes an intricate assembly of protein pigments,featuring extrinsic and intrinsic polypeptides within the photosynthetic membrane.The low-molecular-weight transmembrane protein Psb...Background Photosystem II(PSII)constitutes an intricate assembly of protein pigments,featuring extrinsic and intrinsic polypeptides within the photosynthetic membrane.The low-molecular-weight transmembrane protein PsbX has been identified in PSII,which is associated with the oxygen-evolving complex.The expression of PsbX gene protein is regulated by light.PsbX’s central role involves the regulation of PSII,facilitating the binding of quinone molecules to the Qb(PsbA)site,and it additionally plays a crucial role in optimizing the efficiency of photosynthesis.Despite these insights,a comprehensive understanding of the PsbX gene’s functions has remained elusive.Results In this study,we identified ten PsbX genes in Gossypium hirsutum L.The phylogenetic analysis results showed that 40 genes from nine species were classified into one clade.The resulting sequence logos exhibited substantial conservation across the N and C terminals at multiple sites among all Gossypium species.Furthermore,the ortholo-gous/paralogous,Ka/Ks ratio revealed that cotton PsbX genes subjected to positive as well as purifying selection pressure might lead to limited divergence,which resulted in the whole genome and segmental duplication.The expression patterns of GhPsbX genes exhibited variations across specific tissues,as indicated by the analysis.Moreover,the expression of GhPsbX genes could potentially be regulated in response to salt,intense light,and drought stresses.Therefore,GhPsbX genes may play a significant role in the modulation of photosynthesis under adverse abiotic conditions.Conclusion We examined the structure and function of PsbX gene family very first by using comparative genom-ics and systems biology approaches in cotton.It seems that PsbX gene family plays a vital role during the growth and development of cotton under stress conditions.Collectively,the results of this study provide basic information to unveil the molecular and physiological function of PsbX genes of cotton plants.展开更多
According to the sequence of the bile salt hydrolase （BSH） gene of Bifidobacterium and the restriction enzyme cutting sites of expression vector pNZ8148, primers were designed and the bile salt hydrolase （BSH） gen...According to the sequence of the bile salt hydrolase （BSH） gene of Bifidobacterium and the restriction enzyme cutting sites of expression vector pNZ8148, primers were designed and the bile salt hydrolase （BSH） gene was gotten from Bacillus bifidus ATCC 29521 by PCR. BSH gene was inserted into lactic acid bacteria expression vector pNZ8148 to construct the recombinant pNZ8148-BSH. The recombinant pNZ8148-BSH was transferred into lactic acid bacteria NZ9000 with electrotransformation method. And the recombinant which could express BSH protein was obtained. It was identified by SDS-PAGE electrophoresis and activity verification. The result could provide a rationale reference for expressing BSH in lactic acid bacteria.展开更多
ObjectiveTo investigate the gene expression changes in normal and degeneration lumbar intervertebral disc in humans, providing information for clinical. MethodsThe PCR products of 4096 human genes were spotted onto a ...ObjectiveTo investigate the gene expression changes in normal and degeneration lumbar intervertebral disc in humans, providing information for clinical. MethodsThe PCR products of 4096 human genes were spotted onto a kind of chemical-material-coated-glass slides. The total RNAs were isolated from the tissues. Both the mRNAs from the degeneration and normal lumbar intervertebral disc in humans were reversely transcribed to the cDNAs, which used as the hybridization probes with the incorporations of fluorescent dUTP. The mixed probes were then hybridized to the cDNA microarray. After high-stringent washing, the cDNA microarray was scanned for the fluorescent signals and analyzed with computer image analysis. ResultsAmong the 4096 targets, there were 706 genes whose expression levels differed between the degeneration and normal lumbar intervertebral disc in all cases, comprising 298 up-regulated and 358 down-regulated ones. ConclusionDNA microarray technology is an effective technique in screening for differently expressed genes between the degeneration and normal lumbar intervertebral disc. Cell apoptosis plays an important role in the process of lumbar intervertebral disc degeneration.展开更多
Methionine and lysine are restrictive essential amino acids of livestock, they are also the most attentive indexes in the feed production to carry out the quality control and quality evaluation. Their contents in feed...Methionine and lysine are restrictive essential amino acids of livestock, they are also the most attentive indexes in the feed production to carry out the quality control and quality evaluation. Their contents in feed directly affect livestock protein synthesis. Bacillus natto has excellent probiotic properties. In this experiment, we used the genetic engineering method, fusion PCR technique, to connect methionine-rich gene （zein） from maize endosperm protein with lysine-rich gene （Cflr） from the pepper anther, then the fusion gene was inserted into the expression vector pHT43, and the recombinant plasmid pHT43/zein-Cflr was constructed. The recombinant plasmid was transferred into Bacillus natto, and induced by IPTG for the expression of the fusion gene. We found an apparent band at 40 ku site for the recombinant strain by SDS-PAGE. The contents of methionine and lysine were individually detected with HPLC, the quantities of methionine and lysine in the recombinant strain increased by 18.37% and 24.68% than the wild one, respectively. We also verified the stability of the recombinant bacterium during passaging, and found the stability was 100%. This study provided research-basis for the application of the recombined Bacillus natto as feed additive.展开更多
We cloned and expressed bile salt hydrolase gene ofLactobacillus plantarum M1-UVS29 in Lactococcus lactis NZ9000 successfully. Gene-specific primers for amplification of L. plantarum bsh were designed by using sequenc...We cloned and expressed bile salt hydrolase gene ofLactobacillus plantarum M1-UVS29 in Lactococcus lactis NZ9000 successfully. Gene-specific primers for amplification of L. plantarum bsh were designed by using sequence which availabled from GenBank. The production of PCR amplicon was confirmed by sequencing and cloned into pMD18-T vector, and then recombined into expression vector pNZ8148 and yielding vector pNZ8148-BSH, pNZ8148-BSH was transferred into Lactococcus lactis NZ9000. Sequencing indicated that the cloned bsh fragment contained 995 nucleotides, and shared 99.3% sequence homology with bsh gene from L. plantarum MBUL10. Cloned bsh fragment was successfully transduced into NICE expression system and confirmed by PCR and restriction digest. Recombinant BSH protein was analyzed by SDS-PAGE. The molecular weight of BSH protein was approximately 37 ku. Activity of the expressed protein was 0.77 μmol· min^-1. The successfully expressed proteins by genetic engineering technology made the function of lactic acid bacteria be abundant and laid the foundation for further researches into cholesterol-lowering lactic acid bacterium food and probiotics.展开更多
Background:INDETERMINATE DOMAIN(IDD)transcription factors form one of the largest and most conserved gene families in plant kingdom and play important roles in various processes of plant growth and development,such as...Background:INDETERMINATE DOMAIN(IDD)transcription factors form one of the largest and most conserved gene families in plant kingdom and play important roles in various processes of plant growth and development,such as flower induction in term of flowering control.Till date,systematic and functional analysis of IDD genes remained infancy in cotton.Results:In this study,we identified total of 162 IDD genes from eight different plant species including 65 IDD genes in Gossypium hirsutum.Phylogenetic analysis divided IDDs genes into seven well distinct groups.The gene structures and conserved motifs of GhIDD genes depicted highly conserved exon-intron and protein motif distribution patterns.Gene duplication analysis revealed that among 142 orthologous gene pairs,54 pairs have been derived by segmental duplication events and four pairs by tandem duplication events.Further,Ka/Ks values of most of orthologous/paralogous gene pairs were less than one suggested the purifying selection pressure during evolution.Spatiotemporal expression pattern by qRT-PCR revealed that most of the investigated GhIDD genes showed higher transcript levels in ovule of seven days post anthesis,and upregulated response under the treatments of multiple abiotic stresses.Conclusions:Evolutionary analysis revealed that IDD gene family was highly conserved in plant during the rapid phase of evolution.Whole genome duplication,segmental as well as tandem duplication significantly contributed to the expansion of IDD gene family in upland cotton.Some distinct genes evolved into special subfamily and indicated potential role in the allotetraploidy Gossypium hisutum evolution and development High transcript levels of GhIDD genes in ovules illustrated their potential roles in seed and fiber development Further,upregulated responses of GhIDD genes under the treatments of various abiotic stresses suggested them as important genetic regulators to improve stress resistance in cotton breeding.展开更多
Gene expression profiling at early stages(0～2 DPA) of fiber development in Gossypium hirsutum identified a number of transcription factors which were down regulated in fiberless mutants relative to wild type controls...Gene expression profiling at early stages(0～2 DPA) of fiber development in Gossypium hirsutum identified a number of transcription factors which were down regulated in fiberless mutants relative to wild type controls and which could play a role in controlling early fiber development.Chief among these was GhMYB25,a Mixta-like MYB gene.Transgenic GhMYB25-silenced cotton展开更多
Background： The SWEET （Sugars will eventually be exported transporters） gene family plays multiple roles in plant physiological activities and development process. It participates in reproductive development and in...Background： The SWEET （Sugars will eventually be exported transporters） gene family plays multiple roles in plant physiological activities and development process. It participates in reproductive development and in the process of sugar transport and absorption, plant senescence and stress responses and plant-pathogen interaction. However, thecomprehensive analysis of SWEET genes has not been reported in cotton. Results： In this study, we identified 22, 31, 55 and 60 SWEETgenes from the sequenced genomes of Gossypium orboreum, G. rairnondii, G. hirsutum and G. borbadense, respectively. Phylogenetic tree analysis showed that the SWEET genes could be divided into four groups, which were further classified into 14 sub-clades. Further analysis of chromosomal location, synteny analysis and gene duplication suggested that the orthologs showed a good collinearity and segmental duplication events played a crucial role in the expansion of the family in cotton. Specific MtN3_slv domains were highly conserved between Arabidopsis and cotton by exon-intron organization and motif analysis. In addition, the expression pattern in different tissues indicated that the duplicated genes in cotton might have acquired new functions as a result of sub-functionalization or neo-functionalization. The expression pattern of SWEET genes showed that the different genes were induced by diverse stresses. The identification and functional analysis of SWEET genes in cotton may provide more candidate genes for genetic modification. Conclusion： SWEET genes were classified into four clades in cotton. The expression patterns suggested that the duplicated genes might have experienced a functional divergence. This work provides insights into the evolution of SWEETgenes and more candidates for specific genetic modification, which will be useful in future research.展开更多
To understand the use of real-time reverse transcription-polymerase chain reaction （real-time RT-PCR） for detecting the relative abundance of mRNA, the expression of a tobacco ferrltin gene （NtFer1） was detected b...To understand the use of real-time reverse transcription-polymerase chain reaction （real-time RT-PCR） for detecting the relative abundance of mRNA, the expression of a tobacco ferrltin gene （NtFer1） was detected by Northern blot and real-time RT-PCR. The results indicated that both of the two methods were able to detect mRNA expression of NtFer1 cleady and similady, namely NtFer1 expression was responsive to iron-ovedoad, and the abundance of NtFer1 mRNA was greatly increased after iron loaded for 6 h. To compare the effect and sensitivity of two methods, results revealed that Northern blot need 30 μg of total RNA and at least 3 days for the total protocol performance, whereas real-time RT-PCR only need 2 μg of total RNA and 1.5 h. The real-time RT-PCR is rather sensitive and effective than Northern blot. Real-time RT-PCR analysis can be used to rapidly detect the relative abundance of mRNA expression instead of Northern blot analysis.展开更多
Background： RING H2 finger E3 ligase （RH2FE3） genes encode cysteine rich proteins that mediate E3 ubiquitin ligase activity and degrade target substrates. The roles of these genes in plant responses to phytohormone...Background： RING H2 finger E3 ligase （RH2FE3） genes encode cysteine rich proteins that mediate E3 ubiquitin ligase activity and degrade target substrates. The roles of these genes in plant responses to phytohormones and abiotic stresses are well documented in various species, but their roles in cotton fiber development are poorly understood. To date, genome wide identification and expression analyses of Gossypium hirsutum RH2FE3 genes have not been reported. Methods： We performed computational identification, structural and phylogenetic analyses, chromosomal distribution analysis and estimated KJKs values of G hirsutum RH2FE3 genes. Orthologous and paralogous gene pairs were identified by all versus all BLASTP searches. We predicted cis regulatory elements and analyzed microarray data sets to generate heatmaps at different development stages. Tissue specific expression in cotton fiber, and hormonal and abiotic stress responses were determined by quantitative real time polymerase chain reaction （qRT PCR） analysis. Results： We investigated 140 G hirsutum, 80 G. orboreum, and evolutionary mechanisms and compared them with orthologs 89 G. roimondii putative RH2FB genes and their in Arobidopsis and rice. A domain based analysis of the G hirsutum RH2FE3 genes predicted conserved signature motifs and gene structures. Chromosomal localization showed the genes were distributed across all G hirsutum chromosomes, and 60 duplication events （4 tandem and 56 segmental duplications） and 98 orthologs were detected, cis elements were detected in the promoter regions of G hirsutum RH2FE3 genes. Microarray data and qRT PCR analyses showed that G hirsutum RH2FE3 genes were strongly correlated with cotton fiber development. Additionally, almost all the （brassinolide, gibberellic acid （GA）, indole 3-acetic acid drought, and salt）. dentified genes were up regulated in response to phytohormones （IAA）, and salicylic acid （SA）） and abiotic stresses （cold, heat, Conclusions： The genome wide identification, comprehensive analysis, and characterization of conserved domains and gene structures, as well as phylogenetic analysis, cis element prediction, and expression profile analysis of G hirsutum RH2FE3 genes and their roles in cotton fiber development and responses to plant hormones and abiotic stresses are reported here for the first time. Our findings will contribute to the genome wide analysis of putative RH2FE3 genes in other species and lay a foundation for future physiological and functional research on G hirsutum RH2FE3 genes.展开更多
Fasciclin-like arabinogalactan proteins(FLAs),a subclass of arabinogalactan proteins(AGPs),are usually involved in cell development in plants.To investigate the expression profiling as well
Background:GASA(Giberellic Acid Stimulated in Arabidopsis)gene family plays a crucial role in the phytohormone signaling pathway,growth and development,and stress responses in plants.Many GASA homologs have been ident...Background:GASA(Giberellic Acid Stimulated in Arabidopsis)gene family plays a crucial role in the phytohormone signaling pathway,growth and development,and stress responses in plants.Many GASA homologs have been identified in various plants.Nevertheless,little is known about these proteins in cotton.Results:In the current study,we identified 19,17,25,33,and 38 GASA genes via genome-wide analyses of Gossypium herbaceum,G.arboreum,G.raimondii,G.barbadense,and G.hirsutum,respectively,and performed comprehensive bioinformatics and expression analyses.According to our results,132 GASA proteins shared similar protein structures and were classified into four groups based on the phylogenetic tree.A synteny analysis suggested that segmental duplication was a key driver in the expansion of the GASA gene family.Meanwhile,the cis-element and protein interaction analyses indicated that GhGASA proteins play a significant role in the hormone responses.Transcriptomic and qRT-PCR(Quantitative real time-polymerase chain reaction)analyses revealed diverse expression profiles of the GhGASA genes in different organs under abiotic stresses,indicating that some GhGASA genes possibly participate in fiber development and abiotic-stress responses.Conclusions:The GASA genes in cotton were systematically identified and analyzed for the first time in this paper,and it suggested that the GASA genes are important to the development and growth of cotton.These results will support future exploration of the functions of GASA genes in cotton.展开更多
Background:Calmodulin(CaM)is one of the most important Ca^(2+)signaling receptors because it regulates diverse physiological and biochemical reactions in plants.CaM functions by interacting with CaM-binding proteins(C...Background:Calmodulin(CaM)is one of the most important Ca^(2+)signaling receptors because it regulates diverse physiological and biochemical reactions in plants.CaM functions by interacting with CaM-binding proteins(CaMBPs)to modulate Ca^(2+)signaling.IQ domain(IQD)proteins are plant-specific CaMBPs that bind to CaM by their specific CaM binding sites.Results:In this study,we identified 102 GhIQD genes in the Gossypium hirsutum L.genome.The GhIQD gene family was classified into four clusters(Ⅰ,Ⅱ,Ⅲ,andⅣ),and we then mapped the GhIQD genes to the G.hirsutum L.chromosomes.Moreover,we found that 100 of the 102 GhIQD genes resulted from segmental duplication events,indicating that segmental duplication is the main force driving GhIQD gene expansion.Gene expression pattern analysis showed that a total of 89 GhIQD genes expressed in the elongation stage and second cell wall biosynthesis stage of the fiber cells,suggesting that GhIQD genes may contribute to fiber cell development in cotton.In addition,we found that 20 selected GhIQD genes were highly expressed in various tissues.Exogenous application of MeJA significantly enhanced the expression levels of GhIQD genes.Conclusions:Our study shows that GhIQD genes are involved in fiber cell development in cotton and are also widely induced by MeJA.Thw results provide bases to systematically characterize the evolution and biological functions of GhIQD genes,as well as clues to breed better cotton varieties in the future.展开更多
The paper presented an implement method for a dynamic gene expression display software based on the GTK. This method established the dynamic presentation system of gene expression which according to gene expression da...The paper presented an implement method for a dynamic gene expression display software based on the GTK. This method established the dynamic presentation system of gene expression which according to gene expression data from gene chip hybridize at different time, adopted a linearity combination model and Pearson correlation coefficient algorithm. The system described the gene expression changes in graphic form, the gene expression changes with time and the changes in characteristics of the gene expression, also the changes in relations of the gene expression and regulation relationships among genes. The system also provided an integrated platform for analysis on gene chips data, especially for the research on the network ofgene regulation.展开更多
Specific primers for the MC1R gene of alpacas(GenBank EU1358800) were designed to amplify the cDNA sequence using RT-PCR to seek variation in the sequence and explore the relationship between the expression level of M...Specific primers for the MC1R gene of alpacas(GenBank EU1358800) were designed to amplify the cDNA sequence using RT-PCR to seek variation in the sequence and explore the relationship between the expression level of MC1R gene and alpaca coat color.The MC1R gene from white alpaca was cloned successfully and sequence analysis verified that the MC1R gene,encoding 317 amino acids,was 1081 bp in length.Compared with the existing sequence in GenBank,sequence identity was 99.9%and 7 mutations were found.Primers,designed from the sequence obtained,were used to assess the relative expression of MC1R in alpacas of different coat color using QRT-PCR and SPSS 13.0 software.Relative expression of MC1R in the skin of brown alpacas was 4.32 times higher than that in white alpacas after normalization with GAPDH(P【0.01),indicating that MC1R expression may be related to coat color of alpacas.展开更多
The outer membrane protein, ompA, ofAeromonas veronii has a role in the virulence of the organism and is a potential candidate for vaccine development. In this study, ompA I ofAeromonas veronii strain WA106 was cloned...The outer membrane protein, ompA, ofAeromonas veronii has a role in the virulence of the organism and is a potential candidate for vaccine development. In this study, ompA I ofAeromonas veronii strain WA106 was cloned and sequenced, then, it was expressed in Escherichia coli BL21. The nucleotide sequence of ompA I gene was 1 023 base pairs （GenBank Accession NO.KC748024）, which showed 100% homology with that of A. veronii （NO.AB290200.1）. This predicted protein was composed of 340 amino acid residues. Its molecular weight was 35.78 ku and isoelectric point was 5.18. The protein was a hydrophilic protein containing alpha helix and random coil with percentage of 35.0% and 49.7%, respectively. The tertiary structure, quaternary structure prediction showed that ompA I protein contained two peptide chains. SDS-PAGE showed that the actual value of the fusion protein was consistent with the expected result. It will facilitate further study of the role of ompA I protein.展开更多
The capsid protein precursor （P1）, which plays a major role for the generation of polypeptides of swine vesicular disease virus （SVDV）, was cloned from SVDV HK/70 strain into the retroviral vector pBABE puro and e...The capsid protein precursor （P1）, which plays a major role for the generation of polypeptides of swine vesicular disease virus （SVDV）, was cloned from SVDV HK/70 strain into the retroviral vector pBABE puro and expressed in the mammalian cell line PK15 through the retroviral expression system. The activity of recombinant protein to induce immune response was evaluated in guinea pigs. IFA and Western Blot were used to detect the recombinant protein expression. The results showed that the recombinant protein could be recognized by SVDV positive serum, and animal test showed SVDV-specific antibodies. All of those results indicate that a retroviral-based vaccine carrying the capsid protein precursor （P1） of SVD is able to be expressed in the eukaryotic cell and elicites strong SVDV-specific immune responses in guinea pigs.展开更多
Salinity resistance and differential gene expression associated with salinity in cotton germplasm were studied,because of the large scale area of salinity in China,and its significant negative effects
基金supported by National Natural Science Foundation of China(32060466)Chinese Academy of Agricultural Sciences。
文摘Background Photosystem II(PSII)constitutes an intricate assembly of protein pigments,featuring extrinsic and intrinsic polypeptides within the photosynthetic membrane.The low-molecular-weight transmembrane protein PsbX has been identified in PSII,which is associated with the oxygen-evolving complex.The expression of PsbX gene protein is regulated by light.PsbX’s central role involves the regulation of PSII,facilitating the binding of quinone molecules to the Qb(PsbA)site,and it additionally plays a crucial role in optimizing the efficiency of photosynthesis.Despite these insights,a comprehensive understanding of the PsbX gene’s functions has remained elusive.Results In this study,we identified ten PsbX genes in Gossypium hirsutum L.The phylogenetic analysis results showed that 40 genes from nine species were classified into one clade.The resulting sequence logos exhibited substantial conservation across the N and C terminals at multiple sites among all Gossypium species.Furthermore,the ortholo-gous/paralogous,Ka/Ks ratio revealed that cotton PsbX genes subjected to positive as well as purifying selection pressure might lead to limited divergence,which resulted in the whole genome and segmental duplication.The expression patterns of GhPsbX genes exhibited variations across specific tissues,as indicated by the analysis.Moreover,the expression of GhPsbX genes could potentially be regulated in response to salt,intense light,and drought stresses.Therefore,GhPsbX genes may play a significant role in the modulation of photosynthesis under adverse abiotic conditions.Conclusion We examined the structure and function of PsbX gene family very first by using comparative genom-ics and systems biology approaches in cotton.It seems that PsbX gene family plays a vital role during the growth and development of cotton under stress conditions.Collectively,the results of this study provide basic information to unveil the molecular and physiological function of PsbX genes of cotton plants.
基金Supported by 863 Projects (2008AA10Z311)National Science and Technology Support Projects (2009BADB9B06)+1 种基金Started Post-doctoral Research Grant of Heilongjiang Province (LBH-Q07023)Harbin Technological Innovation of Special Funds (2007RFQXN020)
文摘According to the sequence of the bile salt hydrolase （BSH） gene of Bifidobacterium and the restriction enzyme cutting sites of expression vector pNZ8148, primers were designed and the bile salt hydrolase （BSH） gene was gotten from Bacillus bifidus ATCC 29521 by PCR. BSH gene was inserted into lactic acid bacteria expression vector pNZ8148 to construct the recombinant pNZ8148-BSH. The recombinant pNZ8148-BSH was transferred into lactic acid bacteria NZ9000 with electrotransformation method. And the recombinant which could express BSH protein was obtained. It was identified by SDS-PAGE electrophoresis and activity verification. The result could provide a rationale reference for expressing BSH in lactic acid bacteria.
文摘ObjectiveTo investigate the gene expression changes in normal and degeneration lumbar intervertebral disc in humans, providing information for clinical. MethodsThe PCR products of 4096 human genes were spotted onto a kind of chemical-material-coated-glass slides. The total RNAs were isolated from the tissues. Both the mRNAs from the degeneration and normal lumbar intervertebral disc in humans were reversely transcribed to the cDNAs, which used as the hybridization probes with the incorporations of fluorescent dUTP. The mixed probes were then hybridized to the cDNA microarray. After high-stringent washing, the cDNA microarray was scanned for the fluorescent signals and analyzed with computer image analysis. ResultsAmong the 4096 targets, there were 706 genes whose expression levels differed between the degeneration and normal lumbar intervertebral disc in all cases, comprising 298 up-regulated and 358 down-regulated ones. ConclusionDNA microarray technology is an effective technique in screening for differently expressed genes between the degeneration and normal lumbar intervertebral disc. Cell apoptosis plays an important role in the process of lumbar intervertebral disc degeneration.
基金Supported by the Funding of High Technology Project of Ministry of Science and Technology of China(863 Project,2013AA102504-03)
文摘Methionine and lysine are restrictive essential amino acids of livestock, they are also the most attentive indexes in the feed production to carry out the quality control and quality evaluation. Their contents in feed directly affect livestock protein synthesis. Bacillus natto has excellent probiotic properties. In this experiment, we used the genetic engineering method, fusion PCR technique, to connect methionine-rich gene （zein） from maize endosperm protein with lysine-rich gene （Cflr） from the pepper anther, then the fusion gene was inserted into the expression vector pHT43, and the recombinant plasmid pHT43/zein-Cflr was constructed. The recombinant plasmid was transferred into Bacillus natto, and induced by IPTG for the expression of the fusion gene. We found an apparent band at 40 ku site for the recombinant strain by SDS-PAGE. The contents of methionine and lysine were individually detected with HPLC, the quantities of methionine and lysine in the recombinant strain increased by 18.37% and 24.68% than the wild one, respectively. We also verified the stability of the recombinant bacterium during passaging, and found the stability was 100%. This study provided research-basis for the application of the recombined Bacillus natto as feed additive.
基金Supported by the National Natural Science Fund Project(31171657)Heilongjiang Province Natural Fund Project(ZD201207)Heilongjiang Province Postdoctoral Special Funds(LBH-Q13133)
文摘We cloned and expressed bile salt hydrolase gene ofLactobacillus plantarum M1-UVS29 in Lactococcus lactis NZ9000 successfully. Gene-specific primers for amplification of L. plantarum bsh were designed by using sequence which availabled from GenBank. The production of PCR amplicon was confirmed by sequencing and cloned into pMD18-T vector, and then recombined into expression vector pNZ8148 and yielding vector pNZ8148-BSH, pNZ8148-BSH was transferred into Lactococcus lactis NZ9000. Sequencing indicated that the cloned bsh fragment contained 995 nucleotides, and shared 99.3% sequence homology with bsh gene from L. plantarum MBUL10. Cloned bsh fragment was successfully transduced into NICE expression system and confirmed by PCR and restriction digest. Recombinant BSH protein was analyzed by SDS-PAGE. The molecular weight of BSH protein was approximately 37 ku. Activity of the expressed protein was 0.77 μmol· min^-1. The successfully expressed proteins by genetic engineering technology made the function of lactic acid bacteria be abundant and laid the foundation for further researches into cholesterol-lowering lactic acid bacterium food and probiotics.
基金supported by the Major Research Plan of National Natural Science Foundation of China(NO.31690093)Creative Research Groups of China(31621005)the Agricultural Science and Technology Innovation Program Cooperation and Innovation Mission(CAAS-XTCX2016)
文摘Background:INDETERMINATE DOMAIN(IDD)transcription factors form one of the largest and most conserved gene families in plant kingdom and play important roles in various processes of plant growth and development,such as flower induction in term of flowering control.Till date,systematic and functional analysis of IDD genes remained infancy in cotton.Results:In this study,we identified total of 162 IDD genes from eight different plant species including 65 IDD genes in Gossypium hirsutum.Phylogenetic analysis divided IDDs genes into seven well distinct groups.The gene structures and conserved motifs of GhIDD genes depicted highly conserved exon-intron and protein motif distribution patterns.Gene duplication analysis revealed that among 142 orthologous gene pairs,54 pairs have been derived by segmental duplication events and four pairs by tandem duplication events.Further,Ka/Ks values of most of orthologous/paralogous gene pairs were less than one suggested the purifying selection pressure during evolution.Spatiotemporal expression pattern by qRT-PCR revealed that most of the investigated GhIDD genes showed higher transcript levels in ovule of seven days post anthesis,and upregulated response under the treatments of multiple abiotic stresses.Conclusions:Evolutionary analysis revealed that IDD gene family was highly conserved in plant during the rapid phase of evolution.Whole genome duplication,segmental as well as tandem duplication significantly contributed to the expansion of IDD gene family in upland cotton.Some distinct genes evolved into special subfamily and indicated potential role in the allotetraploidy Gossypium hisutum evolution and development High transcript levels of GhIDD genes in ovules illustrated their potential roles in seed and fiber development Further,upregulated responses of GhIDD genes under the treatments of various abiotic stresses suggested them as important genetic regulators to improve stress resistance in cotton breeding.
文摘Gene expression profiling at early stages(0～2 DPA) of fiber development in Gossypium hirsutum identified a number of transcription factors which were down regulated in fiberless mutants relative to wild type controls and which could play a role in controlling early fiber development.Chief among these was GhMYB25,a Mixta-like MYB gene.Transgenic GhMYB25-silenced cotton
基金supported by the The National Key ResearchDevelopment Program of China(2016YFD0101400,2017YFD0101600)
文摘Background： The SWEET （Sugars will eventually be exported transporters） gene family plays multiple roles in plant physiological activities and development process. It participates in reproductive development and in the process of sugar transport and absorption, plant senescence and stress responses and plant-pathogen interaction. However, thecomprehensive analysis of SWEET genes has not been reported in cotton. Results： In this study, we identified 22, 31, 55 and 60 SWEETgenes from the sequenced genomes of Gossypium orboreum, G. rairnondii, G. hirsutum and G. borbadense, respectively. Phylogenetic tree analysis showed that the SWEET genes could be divided into four groups, which were further classified into 14 sub-clades. Further analysis of chromosomal location, synteny analysis and gene duplication suggested that the orthologs showed a good collinearity and segmental duplication events played a crucial role in the expansion of the family in cotton. Specific MtN3_slv domains were highly conserved between Arabidopsis and cotton by exon-intron organization and motif analysis. In addition, the expression pattern in different tissues indicated that the duplicated genes in cotton might have acquired new functions as a result of sub-functionalization or neo-functionalization. The expression pattern of SWEET genes showed that the different genes were induced by diverse stresses. The identification and functional analysis of SWEET genes in cotton may provide more candidate genes for genetic modification. Conclusion： SWEET genes were classified into four clades in cotton. The expression patterns suggested that the duplicated genes might have experienced a functional divergence. This work provides insights into the evolution of SWEETgenes and more candidates for specific genetic modification, which will be useful in future research.
基金Supported in Part by the Key Project of Chinese Ministry of Education (106065) Heilongjiang Provincial Natural ScienceFoundation (C200533)
文摘To understand the use of real-time reverse transcription-polymerase chain reaction （real-time RT-PCR） for detecting the relative abundance of mRNA, the expression of a tobacco ferrltin gene （NtFer1） was detected by Northern blot and real-time RT-PCR. The results indicated that both of the two methods were able to detect mRNA expression of NtFer1 cleady and similady, namely NtFer1 expression was responsive to iron-ovedoad, and the abundance of NtFer1 mRNA was greatly increased after iron loaded for 6 h. To compare the effect and sensitivity of two methods, results revealed that Northern blot need 30 μg of total RNA and at least 3 days for the total protocol performance, whereas real-time RT-PCR only need 2 μg of total RNA and 1.5 h. The real-time RT-PCR is rather sensitive and effective than Northern blot. Real-time RT-PCR analysis can be used to rapidly detect the relative abundance of mRNA expression instead of Northern blot analysis.
基金supported by the Major Research Plan of National Natural Science Foundation of China(NO.31690093)Young Elite Scientist Sponsorship Program by CAST(China Association for Science and Technology)
文摘Background： RING H2 finger E3 ligase （RH2FE3） genes encode cysteine rich proteins that mediate E3 ubiquitin ligase activity and degrade target substrates. The roles of these genes in plant responses to phytohormones and abiotic stresses are well documented in various species, but their roles in cotton fiber development are poorly understood. To date, genome wide identification and expression analyses of Gossypium hirsutum RH2FE3 genes have not been reported. Methods： We performed computational identification, structural and phylogenetic analyses, chromosomal distribution analysis and estimated KJKs values of G hirsutum RH2FE3 genes. Orthologous and paralogous gene pairs were identified by all versus all BLASTP searches. We predicted cis regulatory elements and analyzed microarray data sets to generate heatmaps at different development stages. Tissue specific expression in cotton fiber, and hormonal and abiotic stress responses were determined by quantitative real time polymerase chain reaction （qRT PCR） analysis. Results： We investigated 140 G hirsutum, 80 G. orboreum, and evolutionary mechanisms and compared them with orthologs 89 G. roimondii putative RH2FB genes and their in Arobidopsis and rice. A domain based analysis of the G hirsutum RH2FE3 genes predicted conserved signature motifs and gene structures. Chromosomal localization showed the genes were distributed across all G hirsutum chromosomes, and 60 duplication events （4 tandem and 56 segmental duplications） and 98 orthologs were detected, cis elements were detected in the promoter regions of G hirsutum RH2FE3 genes. Microarray data and qRT PCR analyses showed that G hirsutum RH2FE3 genes were strongly correlated with cotton fiber development. Additionally, almost all the （brassinolide, gibberellic acid （GA）, indole 3-acetic acid drought, and salt）. dentified genes were up regulated in response to phytohormones （IAA）, and salicylic acid （SA）） and abiotic stresses （cold, heat, Conclusions： The genome wide identification, comprehensive analysis, and characterization of conserved domains and gene structures, as well as phylogenetic analysis, cis element prediction, and expression profile analysis of G hirsutum RH2FE3 genes and their roles in cotton fiber development and responses to plant hormones and abiotic stresses are reported here for the first time. Our findings will contribute to the genome wide analysis of putative RH2FE3 genes in other species and lay a foundation for future physiological and functional research on G hirsutum RH2FE3 genes.
文摘Fasciclin-like arabinogalactan proteins(FLAs),a subclass of arabinogalactan proteins(AGPs),are usually involved in cell development in plants.To investigate the expression profiling as well
基金the National Natural Science Foundation of China(grant no.31701474).
文摘Background:GASA(Giberellic Acid Stimulated in Arabidopsis)gene family plays a crucial role in the phytohormone signaling pathway,growth and development,and stress responses in plants.Many GASA homologs have been identified in various plants.Nevertheless,little is known about these proteins in cotton.Results:In the current study,we identified 19,17,25,33,and 38 GASA genes via genome-wide analyses of Gossypium herbaceum,G.arboreum,G.raimondii,G.barbadense,and G.hirsutum,respectively,and performed comprehensive bioinformatics and expression analyses.According to our results,132 GASA proteins shared similar protein structures and were classified into four groups based on the phylogenetic tree.A synteny analysis suggested that segmental duplication was a key driver in the expansion of the GASA gene family.Meanwhile,the cis-element and protein interaction analyses indicated that GhGASA proteins play a significant role in the hormone responses.Transcriptomic and qRT-PCR(Quantitative real time-polymerase chain reaction)analyses revealed diverse expression profiles of the GhGASA genes in different organs under abiotic stresses,indicating that some GhGASA genes possibly participate in fiber development and abiotic-stress responses.Conclusions:The GASA genes in cotton were systematically identified and analyzed for the first time in this paper,and it suggested that the GASA genes are important to the development and growth of cotton.These results will support future exploration of the functions of GASA genes in cotton.
基金the State Key Laboratory of Cotton Biology Open Fund(grant numbers CB2019A03 and CB2018A07)comprehensive Scientific research fund project of Xianyang Normal University(XSYK20002)+2 种基金the Innovation and Entrepreneurship Training Program for College Students in Shaanxi Province(S202010722071)the National Natural Science Foundation of China(grant number 31872175)Key Research and Development Program of Shaanxi Province(grant number 2019NY-103).
文摘Background:Calmodulin(CaM)is one of the most important Ca^(2+)signaling receptors because it regulates diverse physiological and biochemical reactions in plants.CaM functions by interacting with CaM-binding proteins(CaMBPs)to modulate Ca^(2+)signaling.IQ domain(IQD)proteins are plant-specific CaMBPs that bind to CaM by their specific CaM binding sites.Results:In this study,we identified 102 GhIQD genes in the Gossypium hirsutum L.genome.The GhIQD gene family was classified into four clusters(Ⅰ,Ⅱ,Ⅲ,andⅣ),and we then mapped the GhIQD genes to the G.hirsutum L.chromosomes.Moreover,we found that 100 of the 102 GhIQD genes resulted from segmental duplication events,indicating that segmental duplication is the main force driving GhIQD gene expansion.Gene expression pattern analysis showed that a total of 89 GhIQD genes expressed in the elongation stage and second cell wall biosynthesis stage of the fiber cells,suggesting that GhIQD genes may contribute to fiber cell development in cotton.In addition,we found that 20 selected GhIQD genes were highly expressed in various tissues.Exogenous application of MeJA significantly enhanced the expression levels of GhIQD genes.Conclusions:Our study shows that GhIQD genes are involved in fiber cell development in cotton and are also widely induced by MeJA.Thw results provide bases to systematically characterize the evolution and biological functions of GhIQD genes,as well as clues to breed better cotton varieties in the future.
文摘The paper presented an implement method for a dynamic gene expression display software based on the GTK. This method established the dynamic presentation system of gene expression which according to gene expression data from gene chip hybridize at different time, adopted a linearity combination model and Pearson correlation coefficient algorithm. The system described the gene expression changes in graphic form, the gene expression changes with time and the changes in characteristics of the gene expression, also the changes in relations of the gene expression and regulation relationships among genes. The system also provided an integrated platform for analysis on gene chips data, especially for the research on the network ofgene regulation.
基金supported by the National Natural Science Foundation of China(No.30501070)Shanxi Natural Science Foundation(No.20041099)President Foundation of Agricultural University of Hebei (BS2007023)
文摘Specific primers for the MC1R gene of alpacas(GenBank EU1358800) were designed to amplify the cDNA sequence using RT-PCR to seek variation in the sequence and explore the relationship between the expression level of MC1R gene and alpaca coat color.The MC1R gene from white alpaca was cloned successfully and sequence analysis verified that the MC1R gene,encoding 317 amino acids,was 1081 bp in length.Compared with the existing sequence in GenBank,sequence identity was 99.9%and 7 mutations were found.Primers,designed from the sequence obtained,were used to assess the relative expression of MC1R in alpacas of different coat color using QRT-PCR and SPSS 13.0 software.Relative expression of MC1R in the skin of brown alpacas was 4.32 times higher than that in white alpacas after normalization with GAPDH(P【0.01),indicating that MC1R expression may be related to coat color of alpacas.
基金Supported by the Science&Technology Department of Sichuan Province(2013FZ0014)the Construction Project of Postgraduate Academic Degree in Southwest University for Nationalities(2015XWD-S071007)
文摘The outer membrane protein, ompA, ofAeromonas veronii has a role in the virulence of the organism and is a potential candidate for vaccine development. In this study, ompA I ofAeromonas veronii strain WA106 was cloned and sequenced, then, it was expressed in Escherichia coli BL21. The nucleotide sequence of ompA I gene was 1 023 base pairs （GenBank Accession NO.KC748024）, which showed 100% homology with that of A. veronii （NO.AB290200.1）. This predicted protein was composed of 340 amino acid residues. Its molecular weight was 35.78 ku and isoelectric point was 5.18. The protein was a hydrophilic protein containing alpha helix and random coil with percentage of 35.0% and 49.7%, respectively. The tertiary structure, quaternary structure prediction showed that ompA I protein contained two peptide chains. SDS-PAGE showed that the actual value of the fusion protein was consistent with the expected result. It will facilitate further study of the role of ompA I protein.
基金Supported by Key Technology R&D Programme (2006BAD06A03)
文摘The capsid protein precursor （P1）, which plays a major role for the generation of polypeptides of swine vesicular disease virus （SVDV）, was cloned from SVDV HK/70 strain into the retroviral vector pBABE puro and expressed in the mammalian cell line PK15 through the retroviral expression system. The activity of recombinant protein to induce immune response was evaluated in guinea pigs. IFA and Western Blot were used to detect the recombinant protein expression. The results showed that the recombinant protein could be recognized by SVDV positive serum, and animal test showed SVDV-specific antibodies. All of those results indicate that a retroviral-based vaccine carrying the capsid protein precursor （P1） of SVD is able to be expressed in the eukaryotic cell and elicites strong SVDV-specific immune responses in guinea pigs.
文摘Salinity resistance and differential gene expression associated with salinity in cotton germplasm were studied,because of the large scale area of salinity in China,and its significant negative effects
