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番木瓜CpTIFY10A-like基因克隆及表达分析 被引量:3
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作者 吕金慧 王府润 陈萍 《南方农业学报》 CAS CSCD 北大核心 2020年第6期1308-1315,共8页
【目的】克隆番木瓜CpTIFY10A-like基因序列,并分析其表达特性,为探索该基因功能和抗番木瓜环斑病毒(PRSV)分子机制提供理论依据。【方法】结合前期研究的抗PRSV转录组数据,利用RACE对上调表达基因Cp-TIFY10A-like进行克隆,获得该基因c... 【目的】克隆番木瓜CpTIFY10A-like基因序列,并分析其表达特性,为探索该基因功能和抗番木瓜环斑病毒(PRSV)分子机制提供理论依据。【方法】结合前期研究的抗PRSV转录组数据,利用RACE对上调表达基因Cp-TIFY10A-like进行克隆,获得该基因cDNA全长序列,分析其编码区序列(CDS)及进行生物信息学分析。利用已鉴定的PRSV对3月龄番木瓜植株进行不同处理,以感染PRSV且出现病症的植株(CK+)、1.0 mL/L禾甲安(CTS-N)灌施感染PRSV且出现病症的植株(B)为处理组,以清水灌施(不添加禾甲安)不感染PRSV的植株(CK-)为对照组,实时荧光定量PCR(qRT-PCR)检测不同处理下PRSV基因和CpTIFY10A-like基因在番木瓜叶片中的表达情况。最后将CpTIFY10A-like基因CDS序列连接至pET28a-sumo载体上构建原核表达载体,转化Rosetta(DE3)感受态细胞,通过不同浓度IPTG诱导蛋白表达,利用SDS-PAGE电泳检测原核表达情况。【结果】CpTIFY10A-like基因全长为1352 bp,CDS序列为822 bp,编码273个氨基酸残基;其编码蛋白理论等电点(pI)9.23,不稳定系数62.97,亲水性平均系数-0.458,属于不稳定的亲水性碱性蛋白,定位在细胞核上,其二级结构以无规则卷曲和α-螺旋为主,同时含有少量的β-折叠和延伸链。CpTIFY10A-like蛋白与酸枣和白梨TIFY蛋白的氨基酸序列相似性较高,均含有特定的TIFY结构域,属于TIFY蛋白家族,三者在系统发育进化树上聚在同一分支,表明亲缘关系较近。不同处理的番木瓜叶片中,PRSV基因的相对表达量排序为CK+(1.02)>B(0.39)>CK-(0),CpTIFY10A-like基因的相对表达量排序为B(53.12)>CK+(1.15)>CK-(1.02),且CpTIFY10A-like基因在经禾甲安处理的感染PRSV番木瓜叶片中相对表达量显著升高(P<0.05,下同),而PRSV基因的相对表达量显著降低。CpTIFY10A-like基因在原核细胞中表达蛋白的分子量与理论分子量(29.36 kD)相符,且不同浓度IPTG诱导下,IPTG浓度越高,蛋白表达量越多,表明该基因在原核细胞中成功表达。【结论】禾甲安可诱导CpTIFY10A-like基因高效表达,进而通过茉莉酸通路起调节作用以抵抗PRSV感染,即CpTIFY10A-like基因在番木瓜抗PRSV过程中发挥重要调控作用。 展开更多
关键词 番木瓜 CpTIFY10a-like基因 生物信息学 原核表达 禾甲安(CTS-N) 番木瓜环斑病毒(PRSV)
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怀玉山三叶青烟草病毒增殖蛋白2A-like基因克隆及其功能分析 被引量:1
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作者 夏瑾华 李丹丹 +4 位作者 李然 刘贤泥 刘子捷 罗冰 欧丽佳 《江苏农业科学》 北大核心 2023年第2期36-43,共8页
通过怀玉山三叶青试管苗转录组数据库筛选得到怀玉山三叶青烟草病毒增殖蛋白2A-like基因的核心片段,利用反转录-聚合酶链反应(RT-PCR)技术克隆怀玉山三叶青烟草病毒增殖蛋白2A-like基因,并对其进行序列分析、亚细胞定位和器官表达分析... 通过怀玉山三叶青试管苗转录组数据库筛选得到怀玉山三叶青烟草病毒增殖蛋白2A-like基因的核心片段,利用反转录-聚合酶链反应(RT-PCR)技术克隆怀玉山三叶青烟草病毒增殖蛋白2A-like基因,并对其进行序列分析、亚细胞定位和器官表达分析。结果表明,怀玉山三叶青烟草病毒增殖蛋白2A-like基因cDNA总长度为828 bp,G+C含量为46.50%;怀玉山三叶青烟草病毒增殖蛋白2A-like由275个氨基酸组成,分子量为30728.97 u,等电点为5.39,为疏水性蛋白;二级结构由α-螺旋(28.36%)、β-片层(20.73%)、无规则卷曲(50.91%)构成;三级结构为单体跨膜蛋白;预测怀玉山三叶青烟草病毒增殖蛋白2A-like主要存在液泡膜中;怀玉山三叶青烟草病毒增殖蛋白2A-like与葡萄烟草病毒增殖蛋白2A(GenBank:XP_034708140.1、XP_002279170.2)的同源性最高,同源性达到86.79%。通过烟草叶片亚细胞定位分析表明,烟草病毒增殖蛋白2A-like定位于细胞质和细胞核中。实时定量PCR结果显示,烟草病毒增殖蛋白2A-like基因在怀玉山三叶青2个栽培种中的表达存在器官特异性,怀玉2号和怀玉1号均在叶中表达量最高。怀玉山三叶青烟草病毒增殖蛋白2A-like具有典型烟草病毒增殖蛋白2A的结构特征,氨基酸序列及核酸序列与同源物种相似度较高,进化上较为保守,且可降低植株的光合作用效率。 展开更多
关键词 怀玉山三叶青 烟草病毒增殖蛋白2a-like 基因克隆 表达分析 亚细胞定位
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辣椒疫霉菌PiAvr3a-like基因的鉴定及功能分析 被引量:2
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作者 许静 张美祥 +2 位作者 叶廷跃 沈丹宇 窦道龙 《植物病理学报》 CAS CSCD 北大核心 2016年第2期160-168,共9页
疫霉菌分泌的RxLR效应子能干扰寄主植物细胞的正常生理代谢和功能。我们通过同源序列搜索在辣椒疫霉菌中鉴定到9个PiAvr3a的同源基因(PiAvr3a-like),同时发现PiAvr3a-like在大豆疫霉菌中也有分布,表明该类基因可能在疫霉菌致病过程中发... 疫霉菌分泌的RxLR效应子能干扰寄主植物细胞的正常生理代谢和功能。我们通过同源序列搜索在辣椒疫霉菌中鉴定到9个PiAvr3a的同源基因(PiAvr3a-like),同时发现PiAvr3a-like在大豆疫霉菌中也有分布,表明该类基因可能在疫霉菌致病过程中发挥重要作用。从辣椒疫霉菌中成功克隆了5个PiAvr3a-like基因,表达模式分析发现PcAvh128/132/134这3个基因在疫霉菌侵染早期诱导表达,暗示它们可能在侵染前期发挥功能。接种实验表明在本氏烟中瞬时表达其中的PcAvh128可以显著促进辣椒疫霉菌的侵染,而PcAvh132活性相反,其他3个基因没有明显活性。这5个基因均不能抑制INF1和效应因子诱导的细胞死亡,也不能诱导可见的过敏反应。上述结果表明PiAvr3a-like基因不同成员之间功能存在分化,这为疫霉菌和寄主的分子互作研究提供了材料和参考。 展开更多
关键词 辣椒疫霉菌 PiAvr3a PiAvr3a-like 毒性 细胞死亡
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麻疯树JcCAULIFLOWER A-Like基因的克隆、原核表达及表达模式分析 被引量:3
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作者 赵亮清 徐刚 《植物生理学报》 CAS CSCD 北大核心 2019年第11期1685-1694,共10页
CAULIFLOWER(CAL)是与花分生组织的形成有关的基因。通过RACE-PCR技术从麻疯树花蕾中成功获得了1个CAL(JcCAL A-Like)基因c DNA的全长序列,并对其原核表达产物进行质谱鉴定,分析了其亚细胞定位情况及其在麻疯树花不同发育时期的表达情... CAULIFLOWER(CAL)是与花分生组织的形成有关的基因。通过RACE-PCR技术从麻疯树花蕾中成功获得了1个CAL(JcCAL A-Like)基因c DNA的全长序列,并对其原核表达产物进行质谱鉴定,分析了其亚细胞定位情况及其在麻疯树花不同发育时期的表达情况。结果表明:JcCAL A-Like cDNA全长1021 bp,开放阅读框长603 bp,编码200个氨基酸,蛋白质的大小约为23.368 kDa,等电点pI为9.63;表达情况提示该基因可能参与了麻疯树雄花单核花粉粒的形成及雌花的胚囊成熟;此外,亚细胞定位分析显示该基因定位于细胞核中,这表明JcCAL A-Like发挥功能的场所是在细胞核。 展开更多
关键词 麻疯树 花发育 JcCAULIFLOWER a-like基因 表达分析 亚细胞定位
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新型杀虫基因cry39A-like和cry40-like的克隆、表达与活性分析
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作者 丰硕 黄国强 +2 位作者 程佳旭 曹伟平 宋健 《中国生物防治学报》 CSCD 北大核心 2024年第6期1406-1413,共8页
韭菜迟眼蕈蚊是农业上重要难防地下害虫,其幼虫对韭菜等多种作物造成严重为害,研究室前期筛选到1株在室内和田间均对韭菜迟眼蕈蚊幼虫具有高毒力的Bt菌株,但菌株的杀虫蛋白类型尚不明确。本研究对Bt菌株进行了全基因组测序,发现cry39A-l... 韭菜迟眼蕈蚊是农业上重要难防地下害虫,其幼虫对韭菜等多种作物造成严重为害,研究室前期筛选到1株在室内和田间均对韭菜迟眼蕈蚊幼虫具有高毒力的Bt菌株,但菌株的杀虫蛋白类型尚不明确。本研究对Bt菌株进行了全基因组测序,发现cry39A-like和cry40-like两个新型杀虫基因。通过设计特异性引物和PCR获得基因的全长序列,利用生物信息学软件开展序列分析,构建pGS21T表达载体,转入大肠杆菌Rosetta(DE3)表达,最后测定表达蛋白对韭菜迟眼蕈蚊幼虫杀虫活性。结果表明,cry39A-like开放阅读框为1986 bp,编码661个氨基酸,与cry39Aa1亲缘关系最近,氨基酸序列相似性为94.10%,属于第三分类等级,Cry39A-like蛋白大小为76 kDa,对韭菜迟眼蕈蚊幼虫具有杀虫活性,LC_(50)为89.42μg/mL;cry40-like开放阅读框为2001 bp,编码666个氨基酸,与Cry40类聚在一支,亲缘关系最近,氨基酸序列相似性最高为70.88%,属于第二分类等级,Cry40-like蛋白大小为74.5 kDa,对韭菜迟眼蕈蚊幼虫具有杀虫活性,LC_(50)为126.83μg/mL。本研究明确了Bt菌株主要杀虫基因,cry39A-like和cry40-like可作为新型杀虫基因资源在韭菜迟眼蕈蚊的生物防治中应用。 展开更多
关键词 苏云金芽胞杆菌 cry39a-like cry40-like 韭菜迟眼蕈蚊 杀虫活性
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LMM5.1 and LMM5.4,two eukaryotic translation elongation factor 1Alike gene family members,negatively affect cell death and disease resistance in rice 被引量:13
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作者 Jiying Zhao Pengcheng Liu +4 位作者 Chunrong Li Yanyan Wang Lequn Guo Guanghuai Jiang Wenxue Zhai 《Journal of Genetics and Genomics》 SCIE CAS CSCD 2017年第2期107-118,共12页
Lesion mimic mutant(LMM) genes, stimulating lesion formation in the absence of pathogens, play significant roles in immune response. In this study, we characterized a rice lesion mimic mutant, lmm5,which displayed l... Lesion mimic mutant(LMM) genes, stimulating lesion formation in the absence of pathogens, play significant roles in immune response. In this study, we characterized a rice lesion mimic mutant, lmm5,which displayed light-dependent spontaneous lesions. Additionally, lmm5 plants exhibited enhanced resistance to all of the tested races of Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae(Xoo) by increasing the expression of defense-related genes and the accumulation of hydrogen peroxide. Genetic analysis showed that the lesion mimic phenotype of lmm5 was controlled by two genes, lmm5.1 and lmm5.4, which were isolated with a map-based cloning strategy. Remarkably, LMM5.1 and LMM5.4 share a 97.4% amino acid sequence identity, and they each encode a eukaryotic translation elongation factor 1A(e EF1A)-like protein. Besides, LMM5.1 and LMM5.4 were expressed in a tissue-specific and an indicaspecific manner, respectively. In addition, high-throughput m RNA sequencing analysis confirmed that the basal immunity was constitutively activated in the lmm5 mutant. Taken together, these results suggest that the homologous e EF1A-like genes, LMM5.1 and LMM5.4, negatively affect cell death and disease resistance in rice. 展开更多
关键词 Lesion mimic eEF1a-like gene Cell death Disease resistance Rice
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Breast cancer among Iraqi female in their fifth decade: a retrospective study
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作者 Dhuha Mohammed Kasib Manwar Abdulelah Al-Naqqash Ahmed Salih Alshewered 《TMR Cancer》 2020年第1期1-8,共8页
Background:Breast cancer is a major public health concern for women around the world.Breast cancer incidence increases with age,and this is an important factor for the management of this disease.The aim of this study ... Background:Breast cancer is a major public health concern for women around the world.Breast cancer incidence increases with age,and this is an important factor for the management of this disease.The aim of this study was to assess the clinical profiles of female Iraqi breast cancer patients in their fifth decade,and to evaluate patients'molecular profiles and 5-year disease-free survival(DFS).Methods:A retrospective study was conducted at Baghdad Oncology Teaching Hospital between 1 January 2012 and 12 December 2016.Of 979 individuals seen,285 were aged between 40 and 49 years.Results:The median age at diagnosis was 45 years.The highest incidence of breast cancer occurred among women in their fifth(40-49 years old)and sixth(50-59 years old)decades of life(29%and 28%,respectively).The most common stages at diagnosis were stages II and III,with 40.8%and 39.5%,respectively.Luminal A-like cancers were seen in 124 women(58.5%)and 29 women(13.7%)presented with triple-negative cancers.De novo metastatic disease comprised just 6.7%of the group studied.The median DFS was 48 months(95%CI:41.4-54.5 months).Conclusions:The highest incidence of breast cancer occurred among women in their fifth and sixth decades of life.The most common diagnosis was stage II breast cancer of the luminal A molecular subtype.HER-2(human epidermal growth factor receptor 2)overexpression and triple-negative were the least common subtypes.The median DFS was 48 months. 展开更多
关键词 Iraqi breast cancer Molecular classification Luminal a-like DFS
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发作间期癫痫样放电(IED)对癫痫患者减停抗癫痫药物(AED)后复发的影响探究 被引量:1
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作者 罗伟汀 《智慧健康》 2020年第1期96-97,100,共3页
目的评价发作间期癫痫样放电(IED)对癫痫患者减停抗癫痫药物(AED)后复发的影响。方法选择2016年6月到2019年6月我院的癫痫患者36例,按照具体的癫痫发作分类将其分成IED组与非IED组,每个小组有18例,分析减停抗癫痫药物(AED)后复发率、复... 目的评价发作间期癫痫样放电(IED)对癫痫患者减停抗癫痫药物(AED)后复发的影响。方法选择2016年6月到2019年6月我院的癫痫患者36例,按照具体的癫痫发作分类将其分成IED组与非IED组,每个小组有18例,分析减停抗癫痫药物(AED)后复发率、复发患者的年龄和性别、病程指标。结果 IED组的复发率为11.1%,非IED组的复发率为55.5%。分析具体的复发患者临床指标特点,IED组的男性为20.0%,女性为80.0%,非IED组的男性为0.0%,女性为100.0%,可见女性的复发率要高于男性。IED组的年龄为(67.7±1.3)岁,非IED组的年龄为(67.8±2.2)岁。IED组病程为(4.5±0.6)年,非IED组的病程为(4.6±0.8)年。在相互对比之后发现IED组的复发率高于非IED组,相互的比较存在差异,P<0.05。复发患者的年龄偏高,性别多为女性,且病程一般较长。结论在癫痫患者实际治疗期间,减停抗癫痫药物后的复发,很容易受到发作间期癫痫放电因素的影响,应合理的进行分析和研究,正确的探索实际情况。 展开更多
关键词 发作间期癫痫样放电(IED) 癫痫 减停抗癫痫药物(AED) 复发
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Compound hot drought events in the Mei-yu region:influences from polar and tropical regions
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作者 Huixin Li Bo Sun +4 位作者 Zhongshi Zhang Huijun Wang Yeran Zhou Jiani Zeng Botao Zhou 《Science Bulletin》 2025年第2期273-282,共10页
In the Mei-yu region,there has been noticeable increase in the occurrence of compound hot drought(CHD)events in recent years.However,the underlying causes of these occurrences remain poorly understood.To address this ... In the Mei-yu region,there has been noticeable increase in the occurrence of compound hot drought(CHD)events in recent years.However,the underlying causes of these occurrences remain poorly understood.To address this knowledge gap,we conducted a comprehensive study utilizing observational datasets,reanalysis datasets,and four numerical experiments to investigate the associated physical mechanisms.Our findings indicated that the prevalence of CHD events in the Mei-yu region is influenced strongly by two key factors:the decline in Barents Sea ice during February and the presence of a La Ni?alike pattern of sea surface temperature(SST)in April.The decline in Barents Sea ice generates an anomalous Rossby wave in the Arctic that propagates southeastward.The La Ni?a-like SST pattern regulates a Rossby wave over western America,propagating along the subtropical jet stream.These two Rossby waves induce northward movement and strengthened intensity of the subtropical westerly jet in East Asia.The local circulation patterns in the Mei-yu region are influenced by the position and intensity of the subtropical jet,leading to downward motion in accordance with the secondary circulation theory for high-altitude jet streams.Consequently,these local circulation patterns might contribute to occurrence of CHD events.Moreover,our analysis revealed that the impact of Barents Sea ice and the La Nina-like SST pattern can explain approximately two-thirds of the mild CHD events in the Mei-yu region,and that the influence of each is relatively independent.This research underscores influences of polartropical systems on climate extremes in eastern Asia. 展开更多
关键词 Compound hot drought events Mei-yu region June-July Barents Sea ice La Niña-like SST Respective and combined influences
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Heavy Precipitation over the Jing–Jin–Ji Region in Early October: What Controls Its Interannual Variability?
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作者 Jing FENG Tim LI +1 位作者 Ji WANG Linna ZHANG 《Journal of Meteorological Research》 SCIE CSCD 2022年第4期586-600,共15页
Heavy regional precipitation(HRP)over Beijing,Tianjin,and Hebei Province(the Jing–Jin–Ji region or JJJ)in early October(1–10 October)is a high-impact climate event because of travel and outdoor activities by except... Heavy regional precipitation(HRP)over Beijing,Tianjin,and Hebei Province(the Jing–Jin–Ji region or JJJ)in early October(1–10 October)is a high-impact climate event because of travel and outdoor activities by exceptionally large population during the Chinese National Day Holidays(CNDH).What causes the year-to-year variation of the HRP during early October is investigated through an observational analysis.It is found that the HRP arises from moisture transport by southerly anomalies to the west of an anomalous low-level anticyclone over the subtropical northwestern Pacific(SNWP).Sensitivity numerical experiments reveal that the low-level anticyclonic anomaly is caused by a dipole heating pattern over tropical western and central Pacific associated with a La Niña-like SST anomaly(SSTA)pattern in the Pacific and by a negative heating anomaly over North Europe.The latter connects the SNWP anticyclone through a Rossby wave train.Anomalous ascent associated with a positive heating anomaly over the tropical western Pacific may strengthen the local Hadley Cell,contributing to maintenance of the low-level anomalous anticyclone over SNWP and extending westwards of the western Pacific subtropical high(WPSH).Therefore,both the tropical Pacific and midlatitude heating signals over North Europe may be potential predictors for HRP forecast in the JJJ region in early October. 展开更多
关键词 heavy regional precipitation(HRP)in Jing–Jin–Ji(JJJ) Beijing Tianjin and Hebei Province water vapor(WV)transport Rossby wave train diabatic heating anomalies La Niña-like SST anomaly
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