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系统性硬化症合并肺间质病变患者外周血单个核细胞全基因组DNA甲基化和转录组表达谱
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作者 谢艳莉 赵洪军 +3 位作者 罗卉 左晓霞 李全贞 刘思佳 《中南大学学报(医学版)》 CAS CSCD 北大核心 2023年第6期829-836,共8页
目的:分析系统性硬化症(systemic sclerosis,SSc)合并肺间质病变(interstitial lung disease,ILD)患者外周血单个核细胞(peripheral blood mononuclear cells,PBMCs)全基因组DNA甲基化和转录组表达谱,并进一步探讨DNA甲基化对Wnt/β-cat... 目的:分析系统性硬化症(systemic sclerosis,SSc)合并肺间质病变(interstitial lung disease,ILD)患者外周血单个核细胞(peripheral blood mononuclear cells,PBMCs)全基因组DNA甲基化和转录组表达谱,并进一步探讨DNA甲基化对Wnt/β-catenin信号通路和趋化因子信号通路的影响。方法:收集19例SSc患者(SSc组)及18例健康人(对照组)外周血PBMCs。SSc患者中有10例合并ILD(SSc合并ILD亚组)、9例未合并ILD(SSc未合并ILD亚组)。采用Illumina 450K甲基化芯片和Illumina HT-12 v4.0基因表达谱芯片分析全基因组DNA甲基化和基因表达水平,研究DNA甲基化对Wnt/β-catenin和趋化因子信号通路的影响。结果:全基因组DNA甲基化分析发现:与SSc未合并ILD亚组比较,SSc合并ILD亚组存在71个高甲基化位点,98个低甲基化位点。转录组分析发现:与SSc未合并ILD亚组相比,SSc合并ILD亚组有164个基因表达上调,191个基因表达下调。SSc组患者PBMCs中Wnt/β-catenin信号通路中有35个低甲基化基因,其中卷曲同源物1(frizzled-1,FZD1)、丝裂原活化蛋白激酶9(mitogen-activated protein kinase 9,MAPK9)、母亲DPP同源物2(mothers against DPP homolog 2,SMAD2)、转录因子7类似物2(transcription factor 7-like 2,TCF7L2)和无翅型MMTV整合位点家族成员5B(wingless-type MMTV integration site family,member 5B,WNT5B)mRNA在SSc组中的表达显著高于对照组,差异均有统计学意义(均P<0.05)。与SSc未合并ILD亚组比较,SSc合并ILD亚组中Dickkopf相关蛋白2(dickkopf homolog 2,DKK2)、FZD1、MAPK9等多个基因的mRNA表达虽上调,但差异均无统计学意义(均P>0.05)。趋化因子信号通路中有38个低甲基化基因,其中β-抑制蛋白1(β-arrestin 1,ARRB1)、C-X-C基序趋化因子配体10(C-X-C motif chemokine ligand 10,CXCL10)、C-X-C基序趋化因子配体16(C-XC motif chemokine ligand 16,CXCL16)、FGR、中性粒细胞胞浆因子1C(neutrophil cytosolic factor 1C,NCF1C)mRNA在SSc组中的表达显著高于对照组,差异均有统计学意义(均P<0.05)。与SSc未合并ILD亚组比较,SSc合并ILD亚组中ARRB1、CXCL10、CXCL16等多个基因的mRNA表达上调,但差异均无统计学意义(均P>0.05)。结论:SSc合并ILD与SSc无ILD患者存在DNA甲基化和转录组表达谱的差异,且SSc患者PBMCs中存在Wnt/β-catenin和趋化因子信号通路多个基因表达上调,这可能与SSc的发病机制有关。 展开更多
关键词 系统性硬化症 肺间质病变 外周血单个核细胞 全基因DNA甲基化 转录组表达 Wnt/βcatenin信号通路 趋化因子信号通路
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淅川乌骨鸡出壳前后胸肌的全转录组表达谱分析
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作者 石建州 于金冉 +4 位作者 王彦伟 王铁军 李娜 刘阳坤 姚伦广 《畜牧与饲料科学》 2023年第5期1-7,共7页
[目的]分析淅川乌骨鸡出壳前后胸肌生长过程中全转录组表达谱的变化。[方法]采集入孵14 d的淅川乌骨鸡鸡胚胸肌样本(记为X-14 d)和刚出壳1 d的雏鸡胸肌样本(记为X-1 d),围绕编码RNA(messenger RNA,mRNA)、非编码RNA(lncRNA、circRNA和mi... [目的]分析淅川乌骨鸡出壳前后胸肌生长过程中全转录组表达谱的变化。[方法]采集入孵14 d的淅川乌骨鸡鸡胚胸肌样本(记为X-14 d)和刚出壳1 d的雏鸡胸肌样本(记为X-1 d),围绕编码RNA(messenger RNA,mRNA)、非编码RNA(lncRNA、circRNA和miRNA)进行真核生物全转录组测序分析。[结果]淅川乌骨鸡胸肌生长过程中大量基因转录组水平发生显著变化,出孵1 d的雏鸡与入孵14 d的鸡胚相比(X-14 d vs X-1 d),共发现3858个差异表达mRNA,包括1054个上调基因和2804个下调基因;371个差异表达lncRNA,包括222个上调基因和149个下调基因;316个差异表达circRNA,包括148个上调基因和168个下调基因;377个差异表达miRNA,包括159个上调基因和218个下调基因;GO富集分析表明,差异表达mRNA参与多个生物学过程,如生物过程调控、刺激反应、定位、正负调节生物过程、生长过程、免疫系统过程等。对差异表达mRNA进行的KEGG通路富集分析表明,黏附连接、肌动蛋白细胞骨架的调节、氨基酸的生物合成、氧化磷酸化、碳代谢、柠檬酸循环(TCA循环)、紧密连接、间隙连接、代谢途径、焦点黏附、糖酵解/糖异生、丙酮酸代谢、黑素原生成、Notch信号通路等多个信号通路被富集。显著差异表达基因主要与淅川乌骨鸡的生长性状、肉质性状、脂质性状、黑色素生成性状、免疫系统等多个生物性状相关。[结论]mRNA、lncRNA、circRNA、miRNA作为淅川乌骨鸡基因转录组的重要组成部分,在淅川乌骨鸡的生长发育中发挥着重要作用。研究结果为解析淅川乌骨鸡胸肌生长发育过程的基因调控机制提供了参考。 展开更多
关键词 淅川乌骨鸡 胸肌 转录组表达 基因测序
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肝细胞新生脂肪生成过程的全转录组表达谱分析 被引量:1
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作者 张丹彤 逯素梅 +2 位作者 高毅男 叶棣文 马万山 《检验医学与临床》 CAS 2023年第3期328-333,共6页
目的探讨体外非酒精性脂肪性肝病(NAFLD)模型脂肪变性过程全转录组基因表达谱的变化。方法以人肝细胞株LO2(HL-7702)为实验材料,油酸(50μg/mL)作用48 h诱导建立肝细胞脂肪变性模型。围绕lncRNA、circRNA、mRNA进行真核生物全转录组测... 目的探讨体外非酒精性脂肪性肝病(NAFLD)模型脂肪变性过程全转录组基因表达谱的变化。方法以人肝细胞株LO2(HL-7702)为实验材料,油酸(50μg/mL)作用48 h诱导建立肝细胞脂肪变性模型。围绕lncRNA、circRNA、mRNA进行真核生物全转录组测序分析。结果测序结果显示,脂肪变过程中大量的基因转录组水平发生显著变化,上调的基因包括1884个mRNA、351个lncRNA、564个circRNA;下调的基因包括1975个mRNA、297个lncRNA、540个circRNA。针对具有表达差异的mRNA进行GO及Pathway富集分析,发现脂肪酸代谢、脂肪酸降解及炎症等多个信号通路被富集。结论lncRNA、circRNA、mRNA作为人类基因转录组的重要组成部分在NAFLD的发生与发展中发挥重要作用,该研究为肝脏新生脂肪生成过程的基因调控机制提供理论指导。 展开更多
关键词 非酒精性脂肪性肝病 新生脂肪生成 转录组表达 基因测序
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保留非全长读段的ISO-seq数据转录组表达分析 被引量:2
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作者 刘学军 瞿锡垚 张礼 《数据采集与处理》 CSCD 北大核心 2019年第4期594-604,共11页
近年来,基于单分子测序技术的ISO-seq数据以其超长读段长度被越来越多地应用于转录组新型异构体预测研究,但目前大多数研究工作只用到全长读段数据,丢失了非全长读段数据中较多有用信息,因而数据没有得到充分利用。针对这一问题,本文在... 近年来,基于单分子测序技术的ISO-seq数据以其超长读段长度被越来越多地应用于转录组新型异构体预测研究,但目前大多数研究工作只用到全长读段数据,丢失了非全长读段数据中较多有用信息,因而数据没有得到充分利用。针对这一问题,本文在保留非全长读段的基础上提出了两个能同时预测异构体结构和计算其表达比例的模型基于狄利克雷采样的异构体探测与预测(Dirichletsampling for isoform detection and prediction,DSIDP)和基于马尔科夫链的异构体探测与预测(Markovchain for isoform detection and predition,MCIDP)。两个模型均从全长读段中建立异构体预测集,并采用全长读段和非全长读段计算异构体表达比例。DSIDP将所有读段比对至异构体预测集,并使用Dirichlet采样解决多源映射问题,MCIDP使用马尔科夫链模拟基因外显子之间的选择性剪切,该模型还能预测出数据中没有全长读段的异构体。本文采用模拟数据和真实数据验证了两个模型的有效性。 展开更多
关键词 PacBio ISO-seq 转录组表达 第三代测序技术 新型异构体检测 多源映射
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改进的RNA-Seq数据转录组表达分析研究 被引量:3
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作者 石新新 刘学军 张礼 《数据采集与处理》 CSCD 北大核心 2015年第5期1028-1035,共8页
基于高通量测序的RNA-Seq(RNA-sequencing)是用于转录组研究的一种新技术,针对该技术在转录组表达分析研究中存在的读段多源映射和读段非均匀分布等难点,提出一个改进的转录组表达研究方法 LDASeqII(Improvement of latent Dirichlet al... 基于高通量测序的RNA-Seq(RNA-sequencing)是用于转录组研究的一种新技术,针对该技术在转录组表达分析研究中存在的读段多源映射和读段非均匀分布等难点,提出一个改进的转录组表达研究方法 LDASeqII(Improvement of latent Dirichlet allocation for sequencing data)。模型利用剪接异构体结构信息对参数进行约束并进行外显子读段数目归一化处理,解决了读段非均匀分布下的多源映射问题。通过引入"伪外显子"和"伪转录本"分别处理接合区读段和噪声读段。将模型应用到真实数据集上,并与原LDASeq(Latent Dirichlet allocation for sequencing data)模型和目前流行的Cufflinks与RSEM(RNA-Seq by expectation maximization)方法进行对比。结果显示,改进方法获得了更为准确的转录本及基因表达水平计算结果。 展开更多
关键词 基因表达 RNA-SEQ 转录组表达 多源映射 非均匀性
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沉默APE-1对肝癌细胞Hep 3B转录组表达谱及TNF信号通路的影响
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作者 孙志鹏 许光中 +3 位作者 阿民布和 樊庆 彭吉润 张能维 《安徽医科大学学报》 CAS 北大核心 2021年第7期1106-1111,共6页
目的探索沉默APE-1对肝癌细胞Hep 3B转录组表达谱及TNF信号通路的影响。方法基于沉默APE-1表达的肝癌Hep 3B稳定细胞株,进行转录组测序。实验组为APE-1沉默表达的细胞,对照组为转染空白质粒的细胞(n=3)。生物信息学分析肝癌细胞差异表... 目的探索沉默APE-1对肝癌细胞Hep 3B转录组表达谱及TNF信号通路的影响。方法基于沉默APE-1表达的肝癌Hep 3B稳定细胞株,进行转录组测序。实验组为APE-1沉默表达的细胞,对照组为转染空白质粒的细胞(n=3)。生物信息学分析肝癌细胞差异表达基因变化及差异基因的GO、KEGG、DO功能富集,蛋白互作网络。针对TNF信号通路关键基因TNFAIP3、MAP2K6,RT-PCR检测其在实验/对照组细胞中表达,收集10对肝癌/癌旁临床样本,免疫组化检测其蛋白表达。基于TCGA数据库中374例肝癌患者数据,分析APE-1和TNFAIP3,APE-1和MAP2K6表达相关性。结果沉默APE-1导致肝癌细胞Hep 3B中84个基因上调,39个基因下调。差异变化前10位的基因为APE-1、CEMIP、MAP2K6、TAF11L11、CNGB1、COL5A3、OTULINL、DNAJC12、FOLR1、ACKR3。RT-PCR验证TNFAIP3、MAP2K6在沉默APE-1的肝癌细胞Hep 3B中低表达。TNFAIP3、MAP2K6在临床肝癌组织样本中高表达,且APE-1和TNFAIP3,APE-1和MAP2K6表达均呈正相关(P<0.001)。结论APE-1是肝癌发生发展中关键的抗肿瘤基因,其可能通过影响TNF信号通路发挥作用。 展开更多
关键词 APE-1 转录组表达 TNF信号通路 TNFAIP3 MAP2K6
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大麦CIPK基因家族鉴定及表达分析
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作者 高姗姗 冯浩 +3 位作者 黄鑫 张鑫月 张泽阳 刘贵河 《现代农业科技》 2024年第4期156-163,共8页
钙调神经磷酸酶B样相互作用蛋白激酶(CIPK)蛋白家族是由Ca^(2+)介导的植物信号通路中的关键蛋白家族,在植物抗逆和生长发育中发挥着关键作用。本文利用生物信息学方法和转录组数据分析法,进行了大麦CIPK基因家族鉴定及表达分析。结果表... 钙调神经磷酸酶B样相互作用蛋白激酶(CIPK)蛋白家族是由Ca^(2+)介导的植物信号通路中的关键蛋白家族,在植物抗逆和生长发育中发挥着关键作用。本文利用生物信息学方法和转录组数据分析法,进行了大麦CIPK基因家族鉴定及表达分析。结果表明,从大麦基因组数据库中鉴定出31个大麦CIPK基因家族成员,命名为HvCIPK1~HvCIPK31,这些成员分为5个亚族。HvCIPKs基因家族成员具有CIPKs典型的N端激酶结构域和C端NAF调节结构域;蛋白质分子量为40302.27~89926.43 kD,为亲水性蛋白;启动子总共包含11种与非生物胁迫、激素调控以及生长发育相关的顺式作用元件;HvCIPKs与Na^(+)和K^(+)转运体、ABA信号通路关键蛋白(SOS1、AKT1和ABL2)存在相互作用关系;HvCIPK1、HvCIPK2、HvCIPK6、HvCIPK9、HvCIPK11与响应盐碱胁迫相关。该研究可为进一步探索大麦HvCIPKs基因家族功能及调控机制提供理论依据。 展开更多
关键词 大麦 CIPK基因家族 生物信息学 转录组表达
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肿瘤坏死因子α预处理人脐带间充质干细胞的生物学特征分析 被引量:5
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作者 陈自力 曹宁 +4 位作者 徐萌 姜岩 冀美超 郑阳阳 杨莉莉 《中国组织工程研究》 CAS 北大核心 2023年第24期3780-3787,共8页
背景:在部分临床前及临床研究中,人脐带间充质干细胞表现出有希望的治疗结果但仍然存在效果有限的问题。因此,通过预处理的方式增强人脐带间充质干细胞的功能为干细胞培养工艺的开发提供新思路。目的:探索炎性因子肿瘤坏死因子α预处理... 背景:在部分临床前及临床研究中,人脐带间充质干细胞表现出有希望的治疗结果但仍然存在效果有限的问题。因此,通过预处理的方式增强人脐带间充质干细胞的功能为干细胞培养工艺的开发提供新思路。目的:探索炎性因子肿瘤坏死因子α预处理人脐带间充质干细胞的生物学特性变化及作用机制。方法:分离获得的人脐带间充质干细胞培养至第6代,进行炎性因子肿瘤坏死因子α预处理,对比未预处理人脐带间充质干细胞和炎性因子预处理后人脐带间充质干细胞的免疫调节能力、抗炎功能、表面标志物、倍增时间、成脂成骨分化能力的变化,此外对比两组的转录组数据,分析具体机制。结果与结论:(1)与未预处理对照组相比,不同质量浓度肿瘤坏死因子α预处理的人脐带间充质干细胞均显著抑制T细胞增殖,促进Treg细胞增殖,其中1 ng/mL肿瘤坏死因子α作用效果最显著;(2)与未预处理对照组相比,不同质量浓度肿瘤坏死因子α预处理的人脐带间充质干细胞在体外炎症模型中释放更低水平的肿瘤坏死因子α和更高水平的白细胞介素10,其抗炎能力显著增强;(3)两组人脐带间充质干细胞的倍增时间、表面标志物表达无显著差异,均具有成骨成脂分化能力;(4)两组人脐带间充质干细胞的转录组变化主要体现在核因子κB信号通路和肿瘤坏死因子α信号通路的活化;(5)上述实验结果表明,肿瘤坏死因子α预处理后人脐带间充质干细胞的免疫调节及抗炎功能显著增强。 展开更多
关键词 脐带间充质干细胞 炎性因子 预处理 免疫调节 抗炎功能 质量属性 转录组表达 差异基因
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棉纤维次生壁加厚期cDNA-AFLP表达谱剖析和转录组图谱构建 被引量:4
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作者 潘玉欣 马骏 +3 位作者 张桂寅 韩改英 王省芬 马峙英 《科学通报》 EI CAS CSCD 北大核心 2007年第12期1425-1429,共5页
棉纤维强度主要由棉纤维次生壁加厚期决定.以陆地棉品种中棉所8号和海岛棉品种Pima90-53建立的包含109个单株的种间杂交F2群体为材料,用cDNA-AFLP对棉纤维发育次生壁加厚期(20~25DPA)基因表达谱进行剖析,并构建了次生壁加厚期转... 棉纤维强度主要由棉纤维次生壁加厚期决定.以陆地棉品种中棉所8号和海岛棉品种Pima90-53建立的包含109个单株的种间杂交F2群体为材料,用cDNA-AFLP对棉纤维发育次生壁加厚期(20~25DPA)基因表达谱进行剖析,并构建了次生壁加厚期转录组图谱.37对AFLP引物组合共检测到138个多态性表达片段(transcript-derived fragments,TDFs),大小在100~722bp之间,其中75个TDFs(53.62%)在亲本和F2单株中均具有多态性.在75个TDFs中,37个和已发表的棉纤维EST同源性较高:9个和已克隆的棉纤维基因,如半胱氨酸蛋白激酶、液泡H^+-焦磷酸激酶、液泡H^+-ATP催化亚基、半乳糖蛋白、受体蛋白激酶、GIA/RGA-赤霉素应答元件和纤维素合成酶等的编码基因同源性高;其他TDFs可能代表棉纤维发育过程中的新基因片段.在75个TDFs中,有46个集中分布在一个连锁群上。 展开更多
关键词 棉花 CDNA-AFLP 表达片段(TDFs)转录图谱 纤维发育
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茄青枯拉尔氏菌Rs-T02在3,4,5-三羟基苯甲酸甲酯作用下的转录组分析 被引量:1
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作者 汪锴豪 邹承武 +2 位作者 袁高庆 林纬 黎起秦 《植物病理学报》 CAS CSCD 北大核心 2019年第2期192-202,共11页
本文利用Illumina Hi Seq测序技术,对在3,4,5-三羟基苯甲酸甲酯(methyl gallate,MG)作用下和对照条件下的茄青枯拉尔氏菌(Ralstonia solanacearum) Rs-T02菌株的转录组进行测序分析,并用GO和KEGG Pathway富集化分析的方法分析差异表达基... 本文利用Illumina Hi Seq测序技术,对在3,4,5-三羟基苯甲酸甲酯(methyl gallate,MG)作用下和对照条件下的茄青枯拉尔氏菌(Ralstonia solanacearum) Rs-T02菌株的转录组进行测序分析,并用GO和KEGG Pathway富集化分析的方法分析差异表达基因(differentially expressed genes,DEGs)的功能和代谢通路,以初步了解MG对R. solanacearum作用的分子机制。结果表明,MG处理组和对照组的差异表达的基因有2 172个,其中1 037个基因上调,1 135个基因下调。随意挑选10个DEGs进行qRT-PCR验证,其基因表达趋势与转录组测序结果一致。通过对差异表达基因的功能和代谢通路分析发现,与细胞结构相关的肽聚糖水解酶、3-磷酸甘油酰基转移酶和UDP-N-乙酰胞壁酰丙氨酸-D-谷氨酸连接酶等蛋白的编码基因上调表达,蛋白YeaQ、外膜蛋白W和外膜蛋白Bam E等蛋白的编码基因下调表达;与能量代谢相关的ATP合成酶结构蛋白、辅酶Q亚基和细胞色素等蛋白的编码基因上调表达,甘油醛-3-磷酸脱氢酶、甘油醛脱氢酶和异柠檬酸裂解酶等蛋白的编码基因下调表达;与致病性相关的gspD、gspE和gspF等基因上调表达,hrpY、hrpB和gspG等基因下调表达;与细菌抗药性相关的氨基酸的氨酰-tRNA连接酶、核糖体RNA小亚单位甲基转移酶G基因和多重药物外排泵亚基AcrA等蛋白的编码基因上调表达;与细菌运动性相关的flg B、flg C和flg D等基因上调表达。推测MG对Rs-T02菌株的抑菌机制可能与MG影响病菌的细胞结构和能量代谢相关基因的差异表达有关;同时,MG影响病菌T3SS和T2SS相关基因的差异表达,从而影响细菌的致病力。此外,3-磷酸甘油酰基转移酶基因、核糖体RNA小亚单位甲基转移酶G和多重药物外排泵亚基AcrA基因等上调表达,可能与病菌抵抗MG的机制有关。 展开更多
关键词 茄青枯拉尔氏菌 3 4 5-三羟基苯甲酸甲酯 转录组表达
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ERH基因对人膀胱癌细胞周期的影响 被引量:1
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作者 周荣升 韩从辉 +5 位作者 陈波 郝林 史振铎 姜波 张治国 庞昆 《中国医药导报》 CAS 2020年第33期4-8,45,F0004,共7页
目的探讨未成熟同源蛋白增强子(ERH)基因影响人膀胱尿路上皮癌(BUC)细胞株T24增殖及凋亡的分子生物学机制。方法将BUC T24细胞分为ERH敲除组和ERH正常组,通过人转录组表达谱芯片技术检测全基因谱中受ERH敲除影响的基因表达情况。通过生... 目的探讨未成熟同源蛋白增强子(ERH)基因影响人膀胱尿路上皮癌(BUC)细胞株T24增殖及凋亡的分子生物学机制。方法将BUC T24细胞分为ERH敲除组和ERH正常组,通过人转录组表达谱芯片技术检测全基因谱中受ERH敲除影响的基因表达情况。通过生物信息分析技术、生物功能富集和蛋白互作分析技术确定ERH基因影响人BUC T24细胞增殖及凋亡的可能分子生物学机制,并通过功能实验加以验证。结果人转录组表达谱芯片结果显示,ERH基因在ERH敲除组表达明显降低,FC值为-4.50,P<0.0001。ERH敲除组比ERH正常组有344个基因表达上调,254个基因表达下调(截断值|FC|>2)。生物功能富集分析结果提示,ERH基因与细胞因子、肿瘤坏死因子、P53下调、细胞凋亡等密切有关;蛋白互作分析技术结果发现ERH基因可能通过细胞周期蛋白影响BUC T24细胞的增殖与凋亡。功能实验提示ERH基因通过细胞周期(主要影响细胞周期的S期)产生上述作用。结论ERH基因通过调控细胞周期影响人膀胱癌T24细胞增殖和凋亡。 展开更多
关键词 未成熟同源蛋白增强子基因 膀胱癌 增殖与凋亡 细胞周期 转录组表达谱芯片技术
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Construction of Recombinant Retroviral Vector Containing HIV-1 Tat Gene and Functional Detection of Expressed Tat in Target Cells 被引量:1
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作者 卢春 钱超 +2 位作者 唐桂霞 黄丽 曾怡 《Journal of Nanjing Medical University》 2003年第6期261-269,共9页
Objective: To construct recombinant retroviral vector containing HIV-1 Tatgene and evaluate the junction of the expressed Tat in target cells. Methods: HIV-1 Tat_(101) genewas recovered from pEV plasmid by Hind Ⅲ dig... Objective: To construct recombinant retroviral vector containing HIV-1 Tatgene and evaluate the junction of the expressed Tat in target cells. Methods: HIV-1 Tat_(101) genewas recovered from pEV plasmid by Hind Ⅲ digestion and cloned into expression plasmid LZESpBMN-Z toconstruct recombinant retroviral expression plasmid named LZRS-Tat_(101). Using the method ofcalcium phosphate, the construct of LZRS-Tat_(101) was then transfected into packaging cell linesPhoenix (ΦNX) which contained env and gal genes encoding structural proteins and pol gene codingfor 3 enzymes ( reverse transcriptase, protease and integrate) essential for retroviral integrationand replication . The stable transfected cell lines was obtained using puromycin to screen for morethan 3 days. Then, immunohistochemical (IHC ) staining was carried out to detect the expressionlevel of Tat_(101) protein in both transiently and stably trancfected ΦNX, respectively. Thesupematants containing recombinant virus collected from transient and stable transfected cells wereemployed to infect 293 cells, respectively, and the expressed Tat in 293 cells was tested by Westernblot. Meantime, the supematants of infected 293 cells was further added to HL3T1 cells which wereHela cell lines containing an HIV-1-LTR/CAT reporter construct to establish a co-culture system.After co-culture for 72 hours, the protein was extracted from HL3T1 cells and used for CAT activityassay. Results: After LZRS- Tat_(101) was transfected into ΦNX, the amount of expressed Tat intransient transfection cells was significantly higher than that in stable transfection cells; Tatcould be detected not only in 293 cells but also in the supematants from 293 cells culture, and Tatin the supematants could activate HIV-1 LTR promoter in HL3T1, resulting in high 'expression of CATlocated at the downstream of LTR. Conclusion: The construct of recombinant retrovirus LZRS-Tat_(101) could express Tat protein in target cells and the expressed Tat was functionally activeand can really exhibit the ability to activate transcription. 展开更多
关键词 HIV-1 tat retroviral expression vector the ability to activatetranscription
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Profiling and Comparison of Color Body Wall Transcriptome of Normal Juvenile Sea Cucumber(Apostichopus japonicus) and Those Produced by Crossing Albino
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作者 MA Deyou YANG Hongsheng SUN Lina 《Journal of Ocean University of China》 SCIE CAS 2014年第6期1033-1042,共10页
Sea cucumber(Apostichopus japonicus) is one of the most important aquaculture animals in China. Usually its normal body color is black that fits its living environment. The juvenile individuals obtained by crossing al... Sea cucumber(Apostichopus japonicus) is one of the most important aquaculture animals in China. Usually its normal body color is black that fits its living environment. The juvenile individuals obtained by crossing albino sea cucumber segregated in body color. To document the transcriptome difference between albino associating sea cucumber and the control, we sequenced their transcriptomes with RNA-seq. Approximately, 4.790 million(M) and 4.884 M reads, 200 nt in length, were generated from the body wall of albino associating sea cucumber and the control, respectively, from them, 9550(46.81%) putative genes were identified. In total, 583 genes were found to express differentially between albino associating sea cucumber and the control. Of these differentially expressed genes(DEGs), 4.8% changed more than five-folds. The expression levels of eight DEGs were confirmed with real-time PCR. The changing trend of these DEGs detected with real-time PCR agreed well with that detected with RNA-seq, although the change degree of some DEGs was different. Four significantly enriched pathways were identified for DEGs, which included phagocytosis, Staphylococcus aureus infection, ECM-receptor interaction and focal adhesion. These pathways were helpful for understanding the physiological difference between albino associating sea cucumber and the control. 展开更多
关键词 Apostichopus japonicus sezparation gene expression profiling RNA-seq real-time PCR
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小鼠前额叶皮层出生后发育过程中篮状细胞的电生理特性和基因表达变化
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作者 朱彦兵 赵冰 +4 位作者 张亚强 王欢 潘雨花蕾 赵羽商 殷东敏 《生理学报》 CAS CSCD 北大核心 2022年第4期525-533,共9页
篮状细胞是一类表达小清蛋白(parvalbumin,PV)的抑制性中间神经元。本文利用增强绿色荧光蛋白(enhanced green fluorescent protein,EGFP)标记篮状细胞的G42转基因小鼠,研究小鼠前额叶皮层出生后发育过程中篮状细胞的电生理特性和基因... 篮状细胞是一类表达小清蛋白(parvalbumin,PV)的抑制性中间神经元。本文利用增强绿色荧光蛋白(enhanced green fluorescent protein,EGFP)标记篮状细胞的G42转基因小鼠,研究小鼠前额叶皮层出生后发育过程中篮状细胞的电生理特性和基因表达变化。麻醉下取出生后第7天(postnatal day 7,P7)、14天(P14)和21天(P21)G42小鼠的前额叶皮层脑组织,制作脑片,人工脑脊液中孵育1 h后利用全细胞膜片钳技术记录篮状细胞的电生理特性;另外,消化上述时间点的脑组织,用流式细胞术分选EGFP标记的篮状细胞,建库后进行转录组测序。从P7到P21,篮状细胞的静息膜电位(resting membrane potential,RMP)超极化程度越来越大,膜输入阻抗(membrane input resistance,Rm)逐渐降低;动作电位(action potential,AP)幅度逐步增大,AP持续时间逐渐缩短,而AP阈电位无显著变化;自发兴奋性突触后电流(spontaneous excitatory postsynaptic currents,sEPSCs)的频率逐渐变大,但幅度无显著变化。转录组测序发现,P14相对于P7的篮状细胞有682个基因的表达水平发生显著改变(22个基因上调,660个基因下调),而P21相对于P14的篮状细胞仅有176个基因的表达水平发生显著改变(107个基因上调,69个基因下调)。这些发育过程中的差异表达基因主要富集在神经凋亡、RNA剪接、高尔基体囊泡转运和轴突导向生长等生物学通路。以上结果揭示了篮状细胞成熟过程中的电生理特性和基因表达变化规律,为进一步研究调控篮状细胞发育的分子机制提供了新的线索。 展开更多
关键词 篮状细胞 静息膜电位 动作电位 兴奋性突触后电位 转录基因表达
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Applications of next-generation sequencing to the study of biological invasions 被引量:5
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作者 Marc RIUS Steve BOURNE +1 位作者 Harry Guy HORNSBY Mark A. CHAPMAN 《Current Zoology》 SCIE CAS CSCD 2015年第3期488-504,共17页
Through the widespread implementation of next-generation sequencing (NGS), analyses of the whole genome (the entire DNA content) and the whole transcriptome (the genes being expressed) are becoming commonplace. ... Through the widespread implementation of next-generation sequencing (NGS), analyses of the whole genome (the entire DNA content) and the whole transcriptome (the genes being expressed) are becoming commonplace. NGS enables the analysis of a vast amount of previously unattainable genetic information. Despite this potential, NGS has yet to be widely imple- mented in genetic studies of biological invasions. The study of the genomic causes and consequences of biological invasions al- lows a deeper understanding of the molecular mechanisms underpinning the invasion process. In this review, we present a brief introduction to NGS followed by a synthesis of current research in the genomics and transcriptomics of adaptation and coloniza- tion. We then highlight research opportunities in the field, including: (1) assembling genomes and transcriptomes of non-model organisms, (2) identifying genomic regions and candidate genes underlying evolutionary processes, and (3) studying the adaptive role of gene expression variation. In particular, because introduced species face a broad range of physiological and biotic chal- lenges when colonizing novel and variable environments, transcriptomics will enable the study of gene regulatory pathways that may be responsible for acclimation or adaptation. To conclude, we identify a number of research approaches that will aid our fu- ture understanding of biological invasions 展开更多
关键词 Exotic species GENOMICS Genotype-environment interactions Invasive species Invasion genetics Invasion route Non-indigenous species Non-native species
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De novo transcriptome assembly of RNA-Seq reads with different strategies 被引量:4
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作者 CHEN Geng YIN KangPing +1 位作者 WANG Charles SHI TieLiu 《Science China(Life Sciences)》 SCIE CAS 2011年第12期1129-1133,共5页
De novo transcriptome assembly is an important approach in RNA-Seq data analysis and it can help us to reconstruct the transcriptome and investigate gene expression profiles without reference genome sequences.We carri... De novo transcriptome assembly is an important approach in RNA-Seq data analysis and it can help us to reconstruct the transcriptome and investigate gene expression profiles without reference genome sequences.We carried out transcriptome assemblies with two RNA-Seq datasets generated from human brain and cell line,respectively.We then determined an efficient way to yield an optimal overall assembly using three different strategies.We first assembled brain and cell line transcriptome using a single k-mer length.Next we tested a range of values of k-mer length and coverage cutoff in assembling.Lastly,we combined the assembled contigs from a range of k values to generate a final assembly.By comparing these assembly results,we found that using only one k-mer value for assembly is not enough to generate good assembly results,but combining the contigs from different k-mer values could yield longer contigs and greatly improve the overall assembly. 展开更多
关键词 RNA-SEQ de novo transcriptome assembly next generation sequencing
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Transcriptome analysis reveals long noncoding RNAs involved in fiber development in cotton (Gossypium arboreum) 被引量:11
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作者 Changsong Zou Qiaolian Wang +6 位作者 Cairui Lu Wencui Yang Youping Zhang Hailiang Cheng Xiaoxu Feng Mtawa Andrew Prosper Guoli Song 《Science China(Life Sciences)》 SCIE CAS CSCD 2016年第2期164-171,共8页
Long noncoding RNAs (lncRNAs) play important roles in various biological regulatory processes in yeast, mammals, and plants. However, no systematic identification of lncRNAs has been reported in Gossypium arboreum. ... Long noncoding RNAs (lncRNAs) play important roles in various biological regulatory processes in yeast, mammals, and plants. However, no systematic identification of lncRNAs has been reported in Gossypium arboreum. In this study, the strand-specific RNA sequencing (ssRNA-seq) of samples from cotton fibers and leaves was performed, and lncRNAs involved in fiber initiation and elongation processes were systematically identified and analyzed. We identified 5,996 lncRNAs, of which 3,510 and 2,486 can be classified as long intergenic noncoding RNAs (lincRNAs) and natural antisense transcripts (IncNAT), respectively. LincRNAs and lncNATs are similar in many aspects, but have some differences in exon number, exon length, and transcript length. Expression analysis revealed that 51.9% of lincRNAs and 54.5% of lncNATs transcripts were preferentially expressed at one stage of fiber development, and were significantly highly expressed than protein-coding tran- scripts (21.7%). During the fiber and rapid elongation stages, rapid and dynamic changes in lncRNAs may contribute to fiber development in cotton. This work describes a set of lncRNAs that are involved in fiber development. The characterization and expression analysis of lncRNAs will facilitate future studies on their roles in fiber development in cotton. 展开更多
关键词 long noncoding RNAs strand specific RNA sequencing fiber TRANSCRIPTOME expression
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Comparative transcriptome analysis of the lichen-forming fungus Endocarpon pusillum elucidates its drought adaptation mechanisms 被引量:6
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作者 WANG YanYan ZHANG XinYu +2 位作者 ZHOU QiMing ZHANG XiaoLing WEI JiangChun 《Science China(Life Sciences)》 SCIE CAS CSCD 2015年第1期89-100,共12页
The lichen-forming fungus was isolated from the desert lichen Endocarpon pusillum that is extremely drought resistant.To understand the molecular mechanisms of drought resistance in the fungus,we employed RNA-seq and ... The lichen-forming fungus was isolated from the desert lichen Endocarpon pusillum that is extremely drought resistant.To understand the molecular mechanisms of drought resistance in the fungus,we employed RNA-seq and quantitative real-time PCR to compare and characterize the differentially expressed genes in pure culture at two different water levels and with that in desiccated lichen.The comparative transcriptome analysis indicated that a total of 1781 genes were differentially expressed between samples cultured under normal and PEG-induced drought stress conditions.Similar to those in drought resistance plants and non-lichenized fungi,the common drought-resistant mechanisms were differentially expressed in E.pusillum.However,the expression change of genes involved in osmotic regulation in E.pusillum is different,which might be the evidence for the feature of drought adaptation.Interestingly,different from other organisms,some genes involved in drought adaption mechanisms showed significantly different expression patterns between the presence and absence of drought stress in E.pusillum.The expression of 23 candidate stress responsive genes was further confirmed by quantitative real-time PCR using dehydrated E.pusillum lichen thalli.This study provides a valuable resource for future research on lichen-forming fungi and shall facilitate future functional studies of the specific genes related to drought resistance. 展开更多
关键词 LICHEN MYCOBIONT DEHYDRATION drought adaption drought resistant
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De novo assembly and comparative analysis of root transcriptomes from different varieties of Panax ginseng C. A. Meyer grown in different environments 被引量:6
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作者 ZHEN Gang ZHANG Lei +7 位作者 DU YaNan YU RenBo LIU XinMin CAO FangRui CHANG Qi DENG Xing Wang XIA Mian HE Hang 《Science China(Life Sciences)》 SCIE CAS CSCD 2015年第11期1099-1110,共12页
Panax ginseng C. A. Meyer is an important traditional herb in eastern Asia. It contains ginsenosides, which are primary bioactive compounds with medicinal properties. Although ginseng has been cultivated since at leas... Panax ginseng C. A. Meyer is an important traditional herb in eastern Asia. It contains ginsenosides, which are primary bioactive compounds with medicinal properties. Although ginseng has been cultivated since at least the Ming dynasty to increase production, cultivated ginseng has lower quantities of ginsenosides and lower disease resistance than ginseng grown under natural conditions. We extracted root RNA from six varieties of fifth-year P. ginseng cultivars representing four different growth conditions, and performed Illumina paired-end sequencing. In total, 163,165,706 raw reads were obtained and used to generate a de novo transcriptome that consisted of 151,763 contigs(76,336 unigenes), of which 100,648 contigs(66.3%) were successfully annotated. Differential expression analysis revealed that most differentially expressed genes(DEGs) were upregulated(246 out of 258, 95.3%) in ginseng grown under natural conditions compared with that grown under artificial conditions. These DEGs were enriched in gene ontology(GO) terms including response to stimuli and localization. In particular, some key ginsenoside biosynthesis-related genes, including HMG-Co A synthase(HMGS), mevalonate kinase(MVK), and squalene epoxidase(SE), were upregulated in wild-grown ginseng. Moreover, a high proportion of disease resistance-related genes were upregulated in wild-grown ginseng. This study is the first transcriptome analysis to compare wild-grown and cultivated ginseng, and identifies genes that may produce higher ginsenoside content and better disease resistance in the wild; these genes may have the potential to improve cultivated ginseng grown in artificial environments. 展开更多
关键词 Panax ginseng de novo assembly paired-end sequencing comparative transcriptome analysis ginsenoside biosynthesis disease resistance genes
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Transcriptome profile of human neuroblastoma cells in the hypomagnetic field 被引量:8
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作者 MO WeiChuan LIU Ying +1 位作者 BARTLETT Perry F HE RongQiao 《Science China(Life Sciences)》 SCIE CAS 2014年第4期448-461,1-3,共14页
Research has shown that the hypomagnetic field(HMF)can affect embryo development,cell proliferation,learning and memory,and in vitro tubulin assembly.In the present study,we aimed to elucidate the molecular mechanism ... Research has shown that the hypomagnetic field(HMF)can affect embryo development,cell proliferation,learning and memory,and in vitro tubulin assembly.In the present study,we aimed to elucidate the molecular mechanism by which the HMF exerts its effect,by comparing the transcriptome profiles of human neuroblastoma cells exposed to either the HMF or the geomagnetic field.A total of 2464 differentially expressed genes(DEGs)were identified,216 of which were up-regulated and2248 of which were down-regulated after exposure to the HMF.These DEGs were found to be significantly clustered into several key processes,namely macromolecule localization,protein transport,RNA processing,and brain function.Seventeen DEGs were verified by real-time quantitative PCR,and the expression levels of nine of these DEGs were measured every 6 h.Most notably,MAPK1 and CRY2,showed significant up-and down-regulation,respectively,during the first 6 h of HMF exposure,which suggests involvement of the MAPK pathway and cryptochrome in the early bio-HMF response.Our results provide insights into the molecular mechanisms underlying the observed biological effects of the HMF. 展开更多
关键词 hypomagnetic field geomagnetic field transcriptome profile massively parallel sequencing MAPK1 CRY2
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