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青杄PwRhomboid基因表达特性及其抗旱作用
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作者 杨沛欣 刘奎 +3 位作者 李兴芬 汪晴晴 曹一博 张凌云 《陕西师范大学学报(自然科学版)》 CAS CSCD 北大核心 2024年第5期1-11,共11页
通过对青杄转录组测序分析,筛选出干旱响应基因PwRhomboid,分析该基因在耐旱方面的功能特性,为揭示青杄的耐旱机理提供理论基础。采用生物信息学对PwRhomboid的氨基酸序列和同源基因进行比对;利用RT-qPCR技术分析PwRhomboid对不同非生... 通过对青杄转录组测序分析,筛选出干旱响应基因PwRhomboid,分析该基因在耐旱方面的功能特性,为揭示青杄的耐旱机理提供理论基础。采用生物信息学对PwRhomboid的氨基酸序列和同源基因进行比对;利用RT-qPCR技术分析PwRhomboid对不同非生物胁迫和激素的响应情况;通过瞬时转化烟草叶片检测PwRhomboid蛋白的亚细胞定位;获得拟南芥和马铃薯的PwRhomboid转基因株系并进行干旱表型验证。研究结果显示,PwRhomboid的氨基酸序列在N端与其同源物种的氨基酸序列具有较大差异,在C端相似度较高,保守性较好;其蛋白主要定位于细胞膜。PwRhomboid的表达量经干旱、低温和脱落酸诱导后发生有显著变化,在成熟叶中表达量最高,其能够提高转基因拟南芥和马铃薯的耐旱性。经干旱处理后,与对照组拟南芥野生型和pCM1205株系相比,PwRhomboid过表达株系PwRhomboid-L1和PwRhomboid-L2的存活率更高,叶绿素荧光最大光化学效率和实际光化学效率更高;在经聚乙二醇模拟干旱处理后,与野生型相比,过表达株系PwRhomboid-L3、PwRhomboid-L4的马铃薯苗高更高。结果表明PwRhomboid的表达量会受到干旱等逆境和激素的影响,过表达PwRhomboid能够提高拟南芥和马铃薯的耐旱性。 展开更多
关键词 青杄 Rhomboid蛋白 耐旱性 基因表达
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Evaluation of the versatility of perforators-enhanced rhomboid flap technique for ischial sores reconstruction
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作者 Waleed Aldabaany Ashraf Hussein Elghamry +1 位作者 Montaser Hosny Mohamed Yassin 《Chinese Journal of Plastic and Reconstructive Surgery》 2024年第2期65-71,共7页
Background: Prolonged pressure on the skin known as pressure sores, can cause frequent injuries to the skin and underlying tissues. However, several prevention approaches, including non-surgical and surgical managemen... Background: Prolonged pressure on the skin known as pressure sores, can cause frequent injuries to the skin and underlying tissues. However, several prevention approaches, including non-surgical and surgical management,are available. In flap surgery for pressure sores, a variety of flap types may be used;each offers certain benefits and some incidences of complications and recurrence. This study evaluated the versatility of the perforatorenhanced rhomboid flap technique for ischial sore construction.Methods: This prospective study was conducted in the Plastic Surgery Department of Fayoum University Hospital.The perforator-enhanced rhomboid flap technique was employed in 20 patients with ischial pressure sores who failed to respond to conservative treatment. A proper history was obtained and a preoperative arterial duplex was performed to determine the site of the prominent perforators for their preservation at the base of the flap.Postoperative follow-ups and evaluations were performed.Results: The mean age of the study group was 43.7 ± 13.1, ranging from 19 to 65 years. Moreover, 80% of the participants were males while 20% were female. The main etiology of the sores was paraplegia in 45% of cases.The mean surface area of sores was 24.8 ± 6.3 cm2. In 75% of cases, the color match of the flap was rated as excellent. Similarly, 70% of the patients rated flap thickness as excellent. In 55% of the cases, the scar appearance was deemed excellent. Overall satisfaction was reported as excellent by 40% of the patients, while only 5% reported poor satisfaction. Approximately, 10% of the cases were identified with seroma, and 30% had wound dehiscence of a mean size of 1.33 ± 0.98 cm and required around 6.8 ± 1.9 weeks to heal.Conclusion: The perforator-enhanced rhomboid flap technique is a versatile method for reconstructing ischial pressure sores owing to its technical ease, short operative time, reliable vascularity, aesthetic outcome, matching skin color, and short recovery time. The main limitations include large defects with large surface areas, lack of a nearby donor site for a rhomboid flap, and scarring from prior procedures around the sore. 展开更多
关键词 Ischial pressure sores Perforators-enhanced rhomboid flaps RECONSTRUCTION
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Effect of layer thickness on the flexural property and microstructure of 3D-printed rhomboid polymer-reinforced cemented tailing composites 被引量:2
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作者 Ziyue Zhao Shuai Cao Erol Yilmaz 《International Journal of Minerals,Metallurgy and Materials》 SCIE EI CAS CSCD 2023年第2期236-249,共14页
For mines with poor ore bodies and surrounding rocks,the general mining method does not allow the ore to be extracted from underground safely and efficiently.For these mines,the downward layered filling mining techniq... For mines with poor ore bodies and surrounding rocks,the general mining method does not allow the ore to be extracted from underground safely and efficiently.For these mines,the downward layered filling mining technique is undoubtedly the most suitable mining method.The downward filling mining technique may eliminate the troubles relating to poor ore deposit conditions,such as production safety,ore loss rate,and depletion rate.However,in this technique,the safety of the artificial roof of the next stratum is of paramount importance.Cementitious tailings backfilling(CTB)that is not sufficiently cemented and causes collapses could threaten ore production.This paper explores a diamond-shaped composite structure to mimic the stability of a glued false roof in an actual infill mine based on the recently emerged three-dimensional(3D)printing technology.Experimental means such as three-point bending and digital image correlation(DIC)techniques were used to explore the flexural characteristics of 3D construction specimens and CTB combinations with different cement/tailings weight ratios at diverse layer heights.The results show that the 3D structure with a 14-mm ply height and CTB has strong flexural characteristics,with a maximum deflection value of 30.1 mm,while the 3D-printed rhomboid polymer(3D-PRP)structure with a 26-mm ply height is slightly worse in terms of flexural strength characteristics,but it has a higher maximum flexural strength of 2.83 MPa.A combination of 3D structure and CTB has more unique mechanical properties than CTB itself.This research work offers practical knowledge on the artificial roof performance of the downward layered filling mining technique and builds a scientific knowledge base regarding the successful application of CTB material in mines. 展开更多
关键词 tailings cemented fills 3D printed rhomboidal polymer three-point bending test digital image correlation
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柔嫩艾美耳球虫rhomboid抗原基因在卡介苗中的克隆与表达 被引量:5
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作者 王秋悦 任科研 +2 位作者 宫鹏涛 李建华 张西臣 《中国预防兽医学报》 CAS CSCD 北大核心 2009年第4期270-274,共5页
以含柔嫩艾美耳球虫rhomboid基因的重组质粒为模板,应用PCR方法扩增该基因完整开放阅读框,克隆至pMD18-T载体中。分别用限制性内切酶PstⅠ/ClaⅠ和PvuⅡ/ClaⅠ进行双酶切,将rhomboid目的基因克隆到大肠杆菌-分支杆菌穿梭表达载体pMV261... 以含柔嫩艾美耳球虫rhomboid基因的重组质粒为模板,应用PCR方法扩增该基因完整开放阅读框,克隆至pMD18-T载体中。分别用限制性内切酶PstⅠ/ClaⅠ和PvuⅡ/ClaⅠ进行双酶切,将rhomboid目的基因克隆到大肠杆菌-分支杆菌穿梭表达载体pMV261和整合表达载体pMV361中,获得重组质粒pRho-261和pRho-361。将重组质粒电穿孔转化卡介苗,经热诱导后对其表达产物进行SDS-PAGE和western blot分析。结果表明成功构建了两个重组质粒,其表达产物与柔嫩艾美耳球虫阳性血清具有免疫反应性,为重组卡介苗在鸡球虫病防治方面的研究奠定了基础。 展开更多
关键词 柔嫩艾美耳球虫 rhomboid基因 rBCG 球虫病疫苗
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斯氏艾美耳球虫Rhomboid基因抗原优势区的筛选与鉴定 被引量:3
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作者 田依凡 赵权 +4 位作者 信彩岩 董井泉 宫鹏涛 李建华 张西臣 《黑龙江畜牧兽医》 CAS 北大核心 2017年第2期156-159,共4页
为了筛选并鉴定斯氏艾美耳球虫Rhomboid基因抗原优势区,试验采用预测斯氏艾美耳球虫Rhomboid基因跨膜结构域,PCR扩增4段目的片段,构建表达载体,并通过SDS-PAGE和Westernblot技术分析目的蛋白。结果表明:成功克隆出4段斯氏艾美耳球虫Rhom... 为了筛选并鉴定斯氏艾美耳球虫Rhomboid基因抗原优势区,试验采用预测斯氏艾美耳球虫Rhomboid基因跨膜结构域,PCR扩增4段目的片段,构建表达载体,并通过SDS-PAGE和Westernblot技术分析目的蛋白。结果表明:成功克隆出4段斯氏艾美耳球虫Rhomboid基因,片段大小分别为102 bp、75 bp、93 bp、99 bp,重组原核表达质粒p ET-32a-Rho1~4构建成功;经SDS-PAGE和Western-blot分析,构建的4段重组质粒都得到了表达且与理论值相符,且重组蛋白中p ET-32aRho4可与兔斯氏艾美耳球虫阳性血清发生特异性反应。 展开更多
关键词 斯氏艾美耳球虫 Rhomboid基因 抗原优势区 原核表达 检测 筛选
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柔嫩艾美耳球虫F2杂交株Rhomboid基因在大肠杆菌中的表达 被引量:2
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作者 张广海 李建华 +2 位作者 宫鹏涛 张西臣 张国才 《吉林农业大学学报》 CAS CSCD 北大核心 2009年第1期93-96,共4页
根据ET-Rhomboid基因开放阅读框,插入有利于表达的酶切位点设计引物,以虫体DNA为模板扩增并克隆了ET-Rhomboid基因,并将该基因与PET-28a(+)质粒载体连接,构建成了原核表达载体PET-ET-Rhom-boid。将构建好的表达质粒转入大肠杆菌DE3,经I... 根据ET-Rhomboid基因开放阅读框,插入有利于表达的酶切位点设计引物,以虫体DNA为模板扩增并克隆了ET-Rhomboid基因,并将该基因与PET-28a(+)质粒载体连接,构建成了原核表达载体PET-ET-Rhom-boid。将构建好的表达质粒转入大肠杆菌DE3,经IPTG诱导表达后,用SDS-PAGE检测表达产物,用蛋白印迹(Western blotting)检测其反应原性。结果表明:含重组质粒的工程菌有明显的表达产物为31 kD的融合蛋白,与推测的融合蛋白的分子量吻合,蛋白印迹检测具有反应原性。 展开更多
关键词 柔嫩艾美耳球虫 Rhomboid基因 原核表达
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OTOPOINT PRESSURE THERAPY FOR TREATMENT OF 32 CASES OF STRAIN OF RHOMBOID MUSCLE
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作者 杨骏 张庆萍 《World Journal of Acupuncture-Moxibustion》 1996年第4期51-53,共3页
We adopted auricular pressure therapy to treat 32 cases with strain of rhomboidmuscle and achieved satisfactory results,namely,the cure rate was 46.87%,improved rate 40.67%,and the total effective rate 87.54%.
关键词 STRAIN of RHOMBOID MUSCLE Otopoint pressure therapy Shoulder-back PAIN
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Contralateral trapezius transfer to treat scapular winging: A case report and review of literature
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作者 Juan José Gil-álvarez Pablo García-Parra +1 位作者 Manuel Anaya-Rojas María del Pilar Martínez-Fuentes 《World Journal of Orthopedics》 2019年第1期33-44,共12页
BACKGROUND No dynamic technique, such as tendon transfer, has been described for scapular winging due to levator scapulae or rhomboid major and minor palsies resulting from an isolated dorsal scapular nerve injury. Th... BACKGROUND No dynamic technique, such as tendon transfer, has been described for scapular winging due to levator scapulae or rhomboid major and minor palsies resulting from an isolated dorsal scapular nerve injury. Thus, we evaluated how the contralateral trapezius compound osteomuscular flap transfer would work in stabilizing lateral scapular winging, and the case is reported here. A literature review was also conducted, and articles relevant to the case are presented.CASE SUMMARY A 37-year-old male patient who had sustained an isolated dorsal scapular nerve injury underwent reconstructive surgery using the contralateral trapezius compound osteomuscular flap transfer technique to treat scapular winging and the consequent pain, and to restore function from the shoulder impairment. As a result, the involved shoulder showed an improved Constant-Murley score, from19.5% to 81.88%.CONCLUSION Contralateral trapezius osteomuscular flap transfer succeeded in stabilizing scapular winging in this case, improving shoulder function and affording pain relief. 展开更多
关键词 TRAPEZOID muscle Osteomuscular flap Scapular winging RHOMBOID muscles NERVE PARALYSIS Dorsal scapular NERVE Case report
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Rhomboid Flap vs. Keystone Perforator Island Flap (KPIF) in the Treatment of Pilonidal Sinus Disease: Comparison of Short-Term Results
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作者 Marius D. Roatis Alexandru V. Georgescu 《International Journal of Clinical Medicine》 2020年第7期454-464,共11页
<b><span style="font-family:Verdana;">Background</span></b><span style="font-family:Verdana;"><strong></strong></span><strong><b><span... <b><span style="font-family:Verdana;">Background</span></b><span style="font-family:Verdana;"><strong></strong></span><strong><b><span style="font-family:Verdana;">:</span></b><span style="font-family:Verdana;"></span></strong><span style="font-family:Verdana;"> Pilonidal disease is a chronic inflammatory disease of the sacrococcygeal region that mainly affects young people. Its incidence is 26 cases per 100,000 persons. Although many techniques have been described, there is no consensus on the treatment of pilonidal sinus disease (PSD). </span><span style="font-family:Verdana;"><strong></strong></span><strong><b><span style="font-family:Verdana;">Materials</span></b><span style="font-family:Verdana;"></span></strong><b><span style="font-family:;" "=""> </span></b><span style="font-family:Verdana;"><strong></strong></span><strong><b><span style="font-family:Verdana;">and</span></b><span style="font-family:Verdana;"></span></strong><span style="font-family:;" "=""> </span><span style="font-family:Verdana;"><strong></strong></span><strong><b><span style="font-family:Verdana;">Methods</span></b><span style="font-family:Verdana;"></span></strong><strong><b><span style="font-family:Verdana;">:</span></b><span style="font-family:Verdana;"></span></strong><b><span style="font-family:;" "=""> </span></b><span style="font-family:Verdana;">This study included 30 patients with PSD who were treated between May 2014 and September 2017. All cases underwent excision and flap reconstruction. The operative time, postoperative complications, the length of hospital stay, painless sitting and walking time, patient satisfaction and recurrence were evaluated prospectively. </span><span style="font-family:Verdana;"><strong></strong></span><strong><b><span style="font-family:Verdana;">Results</span></b><span style="font-family:Verdana;"></span></strong><strong><b><span style="font-family:Verdana;">:</span></b><span style="font-family:Verdana;"></span></strong><b><span style="font-family:;" "=""> </span></b><span style="font-family:Verdana;">The results of this prospective, randomized and comparative study are based on experience of a single surgical centre. All patients were followed up 18 months after discharge from the hospital. There is a difference in surgery durations (minutes) between the two groups (33.86 ± 2.89 min. in “keystone” flap vs. 41.26 ± 4.19 in the “rhombic” flap group) (p = 0.001). There were no significant differences in the length of hospital stay, painless sitting and walking time or patient satisfaction. The total complication rate was 66.6% after rhomboid flap compared with 6.6% after keystone flap. There was no flap necrosis. </span><span style="font-family:Verdana;"><strong></strong></span><strong><b><span style="font-family:Verdana;">Conclusions</span></b><span style="font-family:Verdana;"></span></strong><strong><b><span style="font-family:Verdana;">:</span></b><span style="font-family:Verdana;"></span></strong><span style="font-family:Verdana;"> Both of these methods have shown to be successful in treatment of PSD. The KPIF is associated with the advantages of very simple design, abundant blood supply from the perforator vessels and lower rate of complication.</span> 展开更多
关键词 Pilonidal Sinus Disease Keystone Perforator Flap Rhomboid Flap Surgical Treatment
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云南建水官厅地区晚泥盆世法门期牙形石带的发现及五指山组的厘定意义
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作者 赵云江 包佳凤 +1 位作者 彭成龙 王晓林 《云南地质》 2022年第1期1-8,共8页
官厅地区以南红河北岸前人划分石炭系无古生物依据,对比为扬子地层区石炭系黄龙组。根据官厅地区石炭系泥晶灰岩中采获晚泥盆世法门期牙形石Palmatolepis rhomboids带,结合其夹基性火山岩及硅质岩、硅质条带灰岩等特征,将其厘定为泥盆... 官厅地区以南红河北岸前人划分石炭系无古生物依据,对比为扬子地层区石炭系黄龙组。根据官厅地区石炭系泥晶灰岩中采获晚泥盆世法门期牙形石Palmatolepis rhomboids带,结合其夹基性火山岩及硅质岩、硅质条带灰岩等特征,将其厘定为泥盆系上统五指山组。Palmatolepis rhomboids带在滇东南地区晚泥盆世地层中属首次发现,也是东南地层区广西境内五指山组的同名牙形石带。其岩石地层、生物地层、沉积环境可与广西上泥盆统“南丹(海槽)型”五指山组对比,具相同构造背景。因此,将其划归东南地层区右江地层分区,解决了官厅地区前人划分石炭系分区归属、年代及沉积环境三个重大地质问题,对划分右江地层分区与扬子地层区上扬子地层分区(黔西南地层分区)界线具有重要的地质意义。 展开更多
关键词 牙形石Palmatolepis rhomboids带 晚泥盆世 五指山组 南丹型 云南建水
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EGFP标记柔嫩艾美耳球虫重组卡介苗的构建及其在鸡体内的分布 被引量:1
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作者 王秋悦 任科研 +2 位作者 李建华 张西臣 宫鹏涛 《中国科学(C辑)》 CSCD 北大核心 2008年第11期1084-1089,共6页
鸡球虫病给禽类养殖业带来重大经济影响.本实验利用DNA重组技术将柔嫩艾美尔球虫保护性抗原基因rhomboid和增强型绿色荧光蛋白EGFP基因串连,将其克隆到BCG中,构建EGFP标记柔嫩艾美耳球虫重组卡介苗pMV361-Rho/EGFP.将rBCG口服免疫6日龄... 鸡球虫病给禽类养殖业带来重大经济影响.本实验利用DNA重组技术将柔嫩艾美尔球虫保护性抗原基因rhomboid和增强型绿色荧光蛋白EGFP基因串连,将其克隆到BCG中,构建EGFP标记柔嫩艾美耳球虫重组卡介苗pMV361-Rho/EGFP.将rBCG口服免疫6日龄雏鸡,一周后剖杀,取肝、脾、肺、肾、盲肠各组织器官制作冰冻切片,激光共聚焦显微镜观察各组织器官rhomboid表达情况.另分别于免疫后7,14,21,28天提取肝、脾、肺、肾、盲肠各组织器官总RNA,实时定量RT-PCR方法检测rhomboid目的基因转录情况,比较各时期组织器官rhomboid基因表达量.结果显示,免疫一周后,用激光共聚焦显微镜观察到在肝、脾、肺、肾、盲肠各组织器官中均有EGFP表达.实时定量RT-PCR结果显示,免疫14天后rhomboid基因表达量达到高峰,随后开始下降,至28天后消失.本研究为重组卡介苗疫苗作为抗球虫疫苗的研究奠定基础. 展开更多
关键词 球虫病 rBCG EGFP rhomboid基因 激光共聚焦显微镜 实时定量RT-PCR
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鸡堆型艾美耳球虫Rhomboid蛋白的原核表达及其多克隆抗体的制备和鉴定 被引量:1
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作者 黄艺华 冯俪 +5 位作者 刘鹏 从培君 黄小丹 张瑞莉 杨贵君 李广兴 《中国兽医科学》 CAS CSCD 北大核心 2016年第4期461-466,共6页
为了研究堆型艾美耳球虫Rhomboid蛋白的生物学功能及其作为潜在免疫抗原蛋白的可能性,根据Gen Bank上柔嫩艾美耳球虫Rhomboid基因序列(DQ323509.1)设计特异性引物,以堆型艾美耳球虫c DNA为模板PCR扩增包含Rhomboid基因ORF的片段。根据... 为了研究堆型艾美耳球虫Rhomboid蛋白的生物学功能及其作为潜在免疫抗原蛋白的可能性,根据Gen Bank上柔嫩艾美耳球虫Rhomboid基因序列(DQ323509.1)设计特异性引物,以堆型艾美耳球虫c DNA为模板PCR扩增包含Rhomboid基因ORF的片段。根据测序结果及抗原表位分析,设计表达区域特异性引物,PCR扩增后连接于p GEX-6P-1原核表达载体成功构建p GEX-Rhomboid表达质粒。将此质粒转入大肠杆菌Rosetta感受态细胞中进行IPTG诱导表达,SDS-PAGE分析显示有36 ku的预期蛋白表达。蛋白纯化复性后免疫新西兰大白兔制备兔抗Rhomboid多克隆抗体。ELISA方法检测该多克隆抗体效价达到216,Western-blot及瞬时转染pc DNA-Rhomboid质粒表明,制备的多克隆抗体与堆型艾美耳球虫子孢子目的蛋白和真核质粒瞬时转染BHK21细胞表达蛋白均具有良好的反应性。本试验结果为进一步研究堆型艾美耳球虫Rhomboid蛋白的生物学活性奠定了基础。 展开更多
关键词 堆型艾美耳球虫 Rhomboid蛋白 原核表达 多克隆抗体
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猪弓形虫重组卡介苗的构建及其免疫保护性 被引量:3
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作者 于艳辉 杨升 +6 位作者 姜鑫 信彩岩 李显赫 王伟荣 李建华 张西臣 宫鹏涛 《中国生物制品学杂志》 CAS CSCD 2018年第7期695-700,707,共7页
目的构建弓形虫Rhomboid基因和猪IL-2基因的重组卡介苗(BCG),并分析其免疫保护效果。方法应用RT-PCR技术分别扩增弓形虫Rhomboid基因和猪IL-2基因,克隆至p MD18-T载体中,双酶切鉴定及测序正确后,将弓形虫Rhomboid基因和猪IL-2基因串联... 目的构建弓形虫Rhomboid基因和猪IL-2基因的重组卡介苗(BCG),并分析其免疫保护效果。方法应用RT-PCR技术分别扩增弓形虫Rhomboid基因和猪IL-2基因,克隆至p MD18-T载体中,双酶切鉴定及测序正确后,将弓形虫Rhomboid基因和猪IL-2基因串联后分别连接至穿梭表达载体pMV261和整合表达载体pMV361中,电转化BCG,制备重组BCG pMV261-IL-2-Rho和pMV361-IL-2-Rho。分别经颈部肌肉注射免疫猪,共3次,每次间隔2周,免疫剂量均为108 CFU/只,采用ELISA法检测免疫前后猪体内抗体(IgG)滴度、Th1型及Th2型细胞因子的变化情况;第3次免疫后2周通过腹腔注射的方式接种弓形虫China I株速殖子1×108个/只进行攻虫试验,观察重组BCG对猪的免疫保护效果。结果成功构建了重组BCG p MV261-IL-2-Rho和p MV361-IL-2-Rho。重组BCG免疫猪后,随着免疫次数的增加,血清中抗体滴度也逐渐升高,且与PBS对照组相比,差异有统计学意义(P<0.01)。3次免疫后,pMV261-IL-2-Rho组血清Th1型细胞因子IL-2、IFNγ和IL-12的表达水平均明显提高,与PBS对照组相比,差异有统计学意义(P<0.05),而pMV361-IL-2-Rho组血清IL-2、IFNγ和IL-12的水平虽略有升高,但与PBS对照组相比,差异无统计学意义(P>0.05);各组在免疫过程中Th2型细胞因子IL-4水平均未发生明显变化。攻虫后各组猪出现精神沉郁,食欲下降,体温升高等症状,耐过发病期后的猪,食欲逐渐恢复,体温渐趋正常,各种症状减轻,且各组猪在攻虫后20 d均无死亡。结论构建的基因重组BCG pMV261-IL-2-Rho对猪弓形虫感染具有一定的免疫保护作用。 展开更多
关键词 弓形虫 Rhomboid基因 IL-2基因 重组卡介苗 免疫保护
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弓形虫Rhomboid-IL-2基因重组卡介苗的构建及其对犬的免疫效果 被引量:4
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作者 罗文武 宫鹏涛 +3 位作者 李建华 杨举 李赫 张西臣 《中国病原生物学杂志》 CSCD 北大核心 2016年第8期710-714,共5页
目的构建弓形虫Rhomboid以及犬白介素2基因重组卡介苗,并观察其对免疫犬的免疫效果。方法将鉴定正确的弓形虫Rhombiod基因和犬IL-2基因一起克隆到大肠埃希菌-分枝杆菌穿梭表达载体和整合表达载体中,构建重组质粒pMV261(361)-IL2-Rho。... 目的构建弓形虫Rhomboid以及犬白介素2基因重组卡介苗,并观察其对免疫犬的免疫效果。方法将鉴定正确的弓形虫Rhombiod基因和犬IL-2基因一起克隆到大肠埃希菌-分枝杆菌穿梭表达载体和整合表达载体中,构建重组质粒pMV261(361)-IL2-Rho。将重组质粒转入卡介苗中,对表达产物进行SDS-PAGE电泳分析及Western blot鉴定。分别用重组卡介苗pMV261(361)-IL2-Rho和pMV261(361)-Rho经皮下免疫犬,检测免疫犬体液及细胞免疫水平变化并观察其存活时间。结果成功构建pMV261(361)-IL2-Rho重组质粒,弓形虫Rhomboid基因在重组卡介苗中稳定表达46ku的目的蛋白,Western blot显示该蛋白能被弓形虫感染鼠血清识别。动物实验表明,rBCGpMV261(361)-IL2-Rho能诱导产生较强的体液免疫和Th1型细胞免疫应答反应,rBCGpMV361-IL2-Rho免疫组感染弓形虫后的存活时间组为(14.40±4.50)d,rBCGpMV261-IL2-Rho免疫组感染弓形虫后的存活时间组为(13.00±4.75)d,较BCG对照组存活时间(8.20±5.45)d有所延长。结论 rBCGpMV261(361)-IL2-Rho蛋白具有免疫原性。该重组卡介苗对犬弓形虫感染有一定保护效果。 展开更多
关键词 弓形虫 Rhomboid基因 白介素2 rBCG
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斯氏艾美耳球虫兰州株Rhomboid基因的克隆及原核表达 被引量:2
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作者 曲晗 宫鹏涛 +4 位作者 张西臣 张国才 杨举 李赫 李建华 《中国生物制品学杂志》 CAS CSCD 2012年第3期266-269,共4页
目的克隆并原核表达斯氏艾美耳球虫(Eimeria stiedai,E.stiedai)兰州株Rhomboid基因。方法通过分析基因保守区设计引物,采用RT-PCR方法与3′RACE技术相结合,扩增获得兔斯氏艾美耳球虫Rhomboid基因cDNA全序列,并对其进行序列分析;将获得... 目的克隆并原核表达斯氏艾美耳球虫(Eimeria stiedai,E.stiedai)兰州株Rhomboid基因。方法通过分析基因保守区设计引物,采用RT-PCR方法与3′RACE技术相结合,扩增获得兔斯氏艾美耳球虫Rhomboid基因cDNA全序列,并对其进行序列分析;将获得的Rhomboid基因克隆至原核表达载体pET-28a中,转化E.coli Rosetta(DE3),IPTG诱导表达,并对表达产物进行SDS-PAGE及Western blot分析。结果 Rhomboid基因开放阅读框全长774 bp,编码257个氨基酸残基,预测蛋白质相对分子质量和等电点分别为28 120和8.64,与鸡柔嫩艾美耳球虫(E.tellan)Rhomboid蛋白氨基酸序列的同源性为84.44%;构建的重组原核表达质粒pET-Rhomboid经双酶切和测序证实构建正确;表达的重组蛋白相对分子质量约为29 000,并可与兔抗E.stiedai阳性血清发生特异性反应。结论成功克隆并表达了Rhomboid基因,为其生物学功能及以其作为斯氏艾美耳球虫疫苗候选基因的研究奠定了基础。 展开更多
关键词 艾美耳球虫 Rhomboid基因 RACE 原核细胞 基因表达
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弓形虫Rhomboid-1蛋白基因的克隆及原核表达 被引量:1
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作者 杨拓 宫鹏涛 +4 位作者 杨举 张国才 李赫 张西臣 李建华 《中国病原生物学杂志》 CSCD 北大核心 2013年第10期903-906,共4页
目的克隆并原核表达刚地弓形虫(Toxoplasma gondii)Rhomboid-1(TgROM1)蛋白。方法收集、纯化弓形虫速殖子,用Trizol法提取总RNA,应用RT-PCR技术扩增TgROM1基因,回收的PCR产物与pMD18-T载体连接,构建重组克隆质粒pMD18-T-TgROM1。将重组... 目的克隆并原核表达刚地弓形虫(Toxoplasma gondii)Rhomboid-1(TgROM1)蛋白。方法收集、纯化弓形虫速殖子,用Trizol法提取总RNA,应用RT-PCR技术扩增TgROM1基因,回收的PCR产物与pMD18-T载体连接,构建重组克隆质粒pMD18-T-TgROM1。将重组克隆质粒亚克隆至原核表达载体pGEX-4T-1中,构建重组表达质粒pGEX-4T-1-TgROM1并转化至Rosetta感受态,用IPTG诱导表达,表达产物进行SDS-PAGE和Western blot分析。结果成功克隆了643bp的TgROM1基因,双酶切鉴定重组表达质粒pGEX-4T-1-TgROM1构建正确。SDS-PAGE检测重组表达质粒表达的TgROM1蛋白分子质量约为48ku,Western blot检测表明该蛋白能被鼠抗弓形虫血清识别。结论成功克隆了弓形虫ROM1基因并原核表达了具有反应原性的重组TgROM1蛋白,为该蛋白的功能研究奠定了基础。 展开更多
关键词 弓形虫 Rhomboid蛋白 克隆 原核表达
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微小隐孢子虫与安氏隐孢子虫Rhomboid基因部分序列的克隆和分析
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作者 吴玲 宫鹏涛 +2 位作者 吴树华 张西臣 李建华 《中国病原生物学杂志》 CSCD 2009年第5期343-346,共4页
目的获得我国微小隐孢子虫长春株与安氏隐孢子虫长春株Rhomboid基因(CPRho和CARho)部分序列并确定与国外分离株相应序列的同源性。方法应用RT-PCR技术扩增微小隐孢子虫长春株与安氏隐孢子虫长春株Rhomboid基因片段,然后将其克隆到pMD18-... 目的获得我国微小隐孢子虫长春株与安氏隐孢子虫长春株Rhomboid基因(CPRho和CARho)部分序列并确定与国外分离株相应序列的同源性。方法应用RT-PCR技术扩增微小隐孢子虫长春株与安氏隐孢子虫长春株Rhomboid基因片段,然后将其克隆到pMD18-T载体中,测得序列应用DNAMAN软件进行序列分析。结果经RT-PCR扩增出微小隐孢子虫长春株与安氏隐孢子虫长春株Rhomboid基因部分序列,大小均为723 bp。经DNA MAN软件分析,表明我国微小隐孢子虫长春株与安氏隐孢子虫长春株Rhomboid核苷酸序列同源性为99.6%,氨基酸同源性为99.2%。二者与GenBank公布的相应序列核苷酸同源性分别为99.9%和99.7%,而氨基酸同源性均为99.6%。结论获得了微小隐孢子虫长春株与安氏隐孢子虫长春株Rhomboid基因部分序列,为继续研究该基因的侵入功能等奠定了基础。 展开更多
关键词 微小隐孢子虫 安氏隐孢子虫 Rhomboid基因 克隆
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贝氏隐孢子虫Rhomboid基因的克隆与生物信息学分析
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作者 薛雪 阳毅敏 +1 位作者 陈学秋 杜爱芳 《中国兽医学报》 CAS CSCD 北大核心 2015年第2期235-241,共7页
将Gen Bank中发表的C.parvum、C.hominis和C.muris的菱形体蛋白(Rhomboid,ROM)基因序列进行比对分析,设计合成该基因保守片段的1对特异性引物,以贝氏隐孢子虫基因组DNA为模板,PCR扩增出CbROM基因保守片段,采用染色体步移技术获得基因片... 将Gen Bank中发表的C.parvum、C.hominis和C.muris的菱形体蛋白(Rhomboid,ROM)基因序列进行比对分析,设计合成该基因保守片段的1对特异性引物,以贝氏隐孢子虫基因组DNA为模板,PCR扩增出CbROM基因保守片段,采用染色体步移技术获得基因片段3'端和5'端核苷酸序列,并对所得序列进行测序、拼接与生物信息学分析。结果显示,CbROM基因ORF长度为1 422 bp,编码473个氨基酸,蛋白质理论相对分子质量为52 700,等电点为8.65,是疏水性蛋白质,含有7个串连的跨膜螺旋区,无信号肽;该基因含有典型的菱形体蛋白相关保守结构域,属于菱形体蛋白酶家族;功能位点分析结果显示,CbROM蛋白序列包含6个N-糖基化位点,2个c AMP、c GMP依赖性蛋白激酶磷酸化位点,8个蛋白激酶C磷酸化位点,7个酪蛋白激酶II磷酸化位点和10个N-豆蔻酰化位点;二级结构中包含α-螺旋38.27%,β-折叠16.7%和无规则卷曲45.03%。本试验获得的贝氏隐孢子虫的Rhomboid基因序列,为该基因功能的进一步研究及其基因工程疫苗的研制奠定基础。 展开更多
关键词 贝氏隐孢子虫 Rhomboid基因 染色体步移 生物信息学分析
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Construction of EGFP-tagged rBCG of E.tenella and distribution in chickens 被引量:2
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作者 WANG QiuYue1,LI JianHua1,ZHANG XiChen1,LIU ChengWu2,CAO LiLi1,REN KeYan3,GONG PengTao1,CAI YaNan11 College of Animal Science and Veterinary Medicine,Jilin University,Changchun 130062,China 2 Shenyang Police Dog Training School,Shenyang 110034,China 3 Veterinary Science Institute of Jilin Province,Changchun 130062,China 《Science China(Life Sciences)》 SCIE CAS 2009年第3期278-283,共6页
Chicken coccidiosis is a major parasitic disease with substantial economic burden to the poultry industry.Enhanced green fluorescent protein(EGFP) tagged recombinant Bacille Calmette-Guerin(rBCG),as a fusion protein w... Chicken coccidiosis is a major parasitic disease with substantial economic burden to the poultry industry.Enhanced green fluorescent protein(EGFP) tagged recombinant Bacille Calmette-Guerin(rBCG),as a fusion protein with coccidian rhomboid antigen was constructed to track rBCG in vivo in chickens in this study.Immunization of chickens with one dose of rBCG pMV361-Rho/EGFP induced humoral immune response.The colonization of rBCG in liver,spleen,lung,kidney and caecum was observed by laser confocal microscopy.Real-time quantitative RT-PCR showed a rise expression level of rhomboid protein on the 7th day and a peak on the 14th day and disappearance on the 28th day after immunization.These results have significant implications for the development of rBCG vaccines against avian coccidiosis. 展开更多
关键词 COCCIDIOSIS rBCG EGFP RHOMBOID laser CONFOCAL microscopy real-time RT-PCR
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Self-assembly of Pyrene-modified Rhomboidal Metalloden- drimers via Directional Metal-ligand Bonding Approach 被引量:1
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作者 Menglan He Qing Han +5 位作者 Jiuming He Quanjie Li Zeper Abliz Hongwei Tan Lin Xu Haibo Yang 《Chinese Journal of Chemistry》 SCIE CAS CSCD 2013年第5期663-672,共10页
The design and self-assembly of pyrene-modified rhomboidal metallodendrimers R1--R6 via directional metal-ligand bonding approach is described. By employing pyrene-containing 120° di-Pt(II) acceptor and appropr... The design and self-assembly of pyrene-modified rhomboidal metallodendrimers R1--R6 via directional metal-ligand bonding approach is described. By employing pyrene-containing 120° di-Pt(II) acceptor and appropri- ate 60° dendritic dipyridyl donors, a variety of [G-1]--[G-3] pyrene-modified rhomboidal metallodendrimers with well-defined shape and size were prepared under mild conditions in high yields. The supramolecular dendrimers were characterized with multinuclear NMR (1H and 31p) and mass spectrometry (CSI-TOF-MS). lsotopically re- solved mass spectrometry data support the existence of the pyrene-modified dendrimers with rhomboidal cavities, and NMR data were consistent with the formation of all ensembles. The shape and size of all rhomboidal den- drimers were investigated with the PM6 semiempirical molecular orbital method. Their primary photochemical properties were studied as well. 展开更多
关键词 SELF-ASSEMBLY metal-ligand bonding RHOMBOID metallodendrimer PYRENE
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