【目的】针对杜梨组织中富含多酚、多糖等次生代谢物质特点,筛选从杜梨不同组织中提取高质量RNA的方法。【方法】以杜梨不同组织(根、茎、叶、花和果)为材料,分别采用Trizol法、改良CTAB法和RNAprep Pure Plant Kit试剂盒法提取总RNA,...【目的】针对杜梨组织中富含多酚、多糖等次生代谢物质特点,筛选从杜梨不同组织中提取高质量RNA的方法。【方法】以杜梨不同组织(根、茎、叶、花和果)为材料,分别采用Trizol法、改良CTAB法和RNAprep Pure Plant Kit试剂盒法提取总RNA,利用琼脂糖凝胶电泳和NanoDrop 2000微量分光光度计对RNA进行质量检测。【结果】3种方法均能提出杜梨不同组织中总RNA,但RNAprep Pure Plant Kit法提取的总RNA电泳条带清晰、完整性较好,根、茎、叶、花和果提取的RNA纯度分别为2.07、2.02、2.01、2.01和2.02;浓度分别为186.2 ng·μL^(-1)、174.5 ng·μL^(-1)、163.6 ng·μL^(-1)、167.3 ng·μL^(-1)和121.1 ng·μL^(-1)。以RNAprep Pure Plant Kit试剂盒法提取的总RNA反转录成cDNA为模板,内参基因PbUBI为目的基因进行RT-PCR扩增,均可获得明亮单一条带。【结论】RNAprep Pure Plant Kit试剂盒法更适用于杜梨不同组织总RNA提取,可为下一步进行基因克隆及功能验证等分子生物学研究奠定基础。展开更多
SHAPE （selective T-hydroxyl acylation analyzed by primer extension） chemistry has been widely used in the prediction of RNA secondary structure at single-nucleotide solution. This study was designed to determine the...SHAPE （selective T-hydroxyl acylation analyzed by primer extension） chemistry has been widely used in the prediction of RNA secondary structure at single-nucleotide solution. This study was designed to determine the optimal concentrations of RNA and modifying reagent in SHAPE analysis to improve the accura- cy of structure prediction. The results showed that at the ratio of N-methylisatuic anhydride （NMIA） （μmol/L） to RNA （0μmol/L） of 13： 20, the resulting concen- tration of full-length RNA was the highest, and the sequencing signal was the strongest. So, this ratio is optimal for the analysis and prediction of RNA secondary structure in SHAPE chemistry.展开更多
牦牛所处环境及生物习性明显不同于普通牛,尤其组织样品获取不易,如何将来之不易的样品长时间高效保存意义重大。为此,本研究以天祝白牦牛及本地黄牛为对象,采集胃肠道、肝脏及肾脏样品,Trizol Kit提取RNA,运用食品药品监督管理局(Food ...牦牛所处环境及生物习性明显不同于普通牛,尤其组织样品获取不易,如何将来之不易的样品长时间高效保存意义重大。为此,本研究以天祝白牦牛及本地黄牛为对象,采集胃肠道、肝脏及肾脏样品,Trizol Kit提取RNA,运用食品药品监督管理局(Food and Drug Administration,FDA)规则及Gel-pro软件分析-80℃保存1个月和23个月的RNA浓度及降解程度,研究RNA的保存与质量检测技术。结果表明,牦牛所有胃组织样品RNA放置23个月后浓度均增加,而黄牛的瘤胃及瓣胃却相反;两个基因型牛种空肠及牦牛回肠组织RNA相对稳定;牦牛和黄牛大肠RNA在保存23个月后浓度存在显著差异(P<0.05)。牦牛及黄牛胃部及肝脏和肾脏RNA在保存23个月后,5S条带较清晰,28S和18S条带均不清晰;胃部、肝脏及肾脏28S∶18S在0.34~0.72,肠道28S∶18S在0.43~2.15。高寒牧区牦牛胃肠道等组织按Trizol提取方法可提取高质量的RNA样品,但常规方法保存23个月的RNA样品均会产生不同程度的降解,降解程度亦存在动物基因型及组织部位的差异性。展开更多
[Objectives] This study was conducted to explore the antifungal activity of wood vinegar produced from apricot shell. [Methods]The minimal inhibitory concentration（ MIC） of wood vinegar against Aspergillus fumigatus...[Objectives] This study was conducted to explore the antifungal activity of wood vinegar produced from apricot shell. [Methods]The minimal inhibitory concentration（ MIC） of wood vinegar against Aspergillus fumigatus was determined on agar plates. Then,the mycelial growth and the morphology of the apical vesicles of A. fumigatus cultured in the medium supplemented with 1/2 MIC wood vinegar were observed to evaluate the effect of wood vinegar on A. fumigatus.RNA-Seq was performed to explore the antifungal mechanism of wood vinegar. [Results] The wood vinegar produced from apricot shell could inhibit the growth of A. fumigatus,with a MIC of 20 μl/ml. In the medium supplemented with 1/2 MIC wood vinegar,the mycelial growth of A. fumigatus was suppressed,some apical vesicles were incompletely developed. The gene ontology（ GO） term enrichment analysis based on RNA-Seq showed that the differentially expressed genes（ DEGs）were enriched in metabolic process,cellular process and single-organism process in the category of biological processes,cell,cell part,membrane,organelle and membrane part in the category of cellular components,catalytic activity and binding in the category of molecular function. The significantly up-regulated DEGs were mostly involved in cell,cell part,organelle,organelle part and macro molecular complex in the category of cellular component,while the significantly down-regulated DEGs were mostly involved in biological process,cellular process and single-organism process in the category of biological process,catalytic activity,binding,transporter activity,and nucleic acid binding transcription factor activity in the category molecular function. Moreover,KEGG（ Kyoto Encyclopedia of Genes and Genomes） pathway analysis showed that the DEG unigenes involved in metabolism were the most,followed by those involved in genetic information processing.[Conclusions]The results indicated that the wood vinegar produced from apricot shell might inhibit the growth of A. fumigatus by interfering with t展开更多
文摘【目的】针对杜梨组织中富含多酚、多糖等次生代谢物质特点,筛选从杜梨不同组织中提取高质量RNA的方法。【方法】以杜梨不同组织(根、茎、叶、花和果)为材料,分别采用Trizol法、改良CTAB法和RNAprep Pure Plant Kit试剂盒法提取总RNA,利用琼脂糖凝胶电泳和NanoDrop 2000微量分光光度计对RNA进行质量检测。【结果】3种方法均能提出杜梨不同组织中总RNA,但RNAprep Pure Plant Kit法提取的总RNA电泳条带清晰、完整性较好,根、茎、叶、花和果提取的RNA纯度分别为2.07、2.02、2.01、2.01和2.02;浓度分别为186.2 ng·μL^(-1)、174.5 ng·μL^(-1)、163.6 ng·μL^(-1)、167.3 ng·μL^(-1)和121.1 ng·μL^(-1)。以RNAprep Pure Plant Kit试剂盒法提取的总RNA反转录成cDNA为模板,内参基因PbUBI为目的基因进行RT-PCR扩增,均可获得明亮单一条带。【结论】RNAprep Pure Plant Kit试剂盒法更适用于杜梨不同组织总RNA提取,可为下一步进行基因克隆及功能验证等分子生物学研究奠定基础。
文摘SHAPE （selective T-hydroxyl acylation analyzed by primer extension） chemistry has been widely used in the prediction of RNA secondary structure at single-nucleotide solution. This study was designed to determine the optimal concentrations of RNA and modifying reagent in SHAPE analysis to improve the accura- cy of structure prediction. The results showed that at the ratio of N-methylisatuic anhydride （NMIA） （μmol/L） to RNA （0μmol/L） of 13： 20, the resulting concen- tration of full-length RNA was the highest, and the sequencing signal was the strongest. So, this ratio is optimal for the analysis and prediction of RNA secondary structure in SHAPE chemistry.
文摘牦牛所处环境及生物习性明显不同于普通牛,尤其组织样品获取不易,如何将来之不易的样品长时间高效保存意义重大。为此,本研究以天祝白牦牛及本地黄牛为对象,采集胃肠道、肝脏及肾脏样品,Trizol Kit提取RNA,运用食品药品监督管理局(Food and Drug Administration,FDA)规则及Gel-pro软件分析-80℃保存1个月和23个月的RNA浓度及降解程度,研究RNA的保存与质量检测技术。结果表明,牦牛所有胃组织样品RNA放置23个月后浓度均增加,而黄牛的瘤胃及瓣胃却相反;两个基因型牛种空肠及牦牛回肠组织RNA相对稳定;牦牛和黄牛大肠RNA在保存23个月后浓度存在显著差异(P<0.05)。牦牛及黄牛胃部及肝脏和肾脏RNA在保存23个月后,5S条带较清晰,28S和18S条带均不清晰;胃部、肝脏及肾脏28S∶18S在0.34~0.72,肠道28S∶18S在0.43~2.15。高寒牧区牦牛胃肠道等组织按Trizol提取方法可提取高质量的RNA样品,但常规方法保存23个月的RNA样品均会产生不同程度的降解,降解程度亦存在动物基因型及组织部位的差异性。
基金Supported by the Key Scientific and Technological Research Project for Universities of Hebei Province(ZD2018306)
文摘[Objectives] This study was conducted to explore the antifungal activity of wood vinegar produced from apricot shell. [Methods]The minimal inhibitory concentration（ MIC） of wood vinegar against Aspergillus fumigatus was determined on agar plates. Then,the mycelial growth and the morphology of the apical vesicles of A. fumigatus cultured in the medium supplemented with 1/2 MIC wood vinegar were observed to evaluate the effect of wood vinegar on A. fumigatus.RNA-Seq was performed to explore the antifungal mechanism of wood vinegar. [Results] The wood vinegar produced from apricot shell could inhibit the growth of A. fumigatus,with a MIC of 20 μl/ml. In the medium supplemented with 1/2 MIC wood vinegar,the mycelial growth of A. fumigatus was suppressed,some apical vesicles were incompletely developed. The gene ontology（ GO） term enrichment analysis based on RNA-Seq showed that the differentially expressed genes（ DEGs）were enriched in metabolic process,cellular process and single-organism process in the category of biological processes,cell,cell part,membrane,organelle and membrane part in the category of cellular components,catalytic activity and binding in the category of molecular function. The significantly up-regulated DEGs were mostly involved in cell,cell part,organelle,organelle part and macro molecular complex in the category of cellular component,while the significantly down-regulated DEGs were mostly involved in biological process,cellular process and single-organism process in the category of biological process,catalytic activity,binding,transporter activity,and nucleic acid binding transcription factor activity in the category molecular function. Moreover,KEGG（ Kyoto Encyclopedia of Genes and Genomes） pathway analysis showed that the DEG unigenes involved in metabolism were the most,followed by those involved in genetic information processing.[Conclusions]The results indicated that the wood vinegar produced from apricot shell might inhibit the growth of A. fumigatus by interfering with t
