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Molecular Biology

分子生物学

基本信息

批准号：
8327070
负责人：
Ruth Margrit Ruprecht
金额：
$ 24.28万
依托单位：
DANA-FARBER CANCER INST
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-05 至 2013-08-16
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8327070
关键词：
Adenovirus Vector Animals Antigens Biological Assay Chimera organism DNA Dana-Farber Cancer Institute Diagnostic Disease Disease Progression Engineering Enrollment Ensure Evolution Funding Generations Goals HIV Humoral Immunities Infant Infection Laboratories Leadership Macaca mulatta Medicine Modeling Molecular Biology Molecular Evolution Monitor Monkeys Mothers Pattern Phylogenetic Analysis Plasma Primates Principal Investigator Procedures Progress Reports Proteins Reagent Recombinants Research Retroviridae Infections Reverse Transcriptase Polymerase Chain Reaction Role SIV Safety Scientist Screening procedure Shuttle Vectors Simian Retroviruses Simian T-lymphotropic virus 1 Switzerland Time Training Universities Vaccination Vaccines Vaccinia virus Vertebral column Viral Load result Viral load measurement Virus Wood material Work Writing Zambia base env Genes experience expression vector instructor medical schools member mutant novel pediatric human immunodeficiency virus programs protein expression protein purification simian human immunodeficiency virus transmission process vector viral RNA

项目摘要

The overall goal of Core A is to provide molecular biology expertise and support for the Program Project by closely interacting with all three Projects and Cores. Core A will construct novel simian-human immunodeficiency virus (SHIV) strains encoding R5 env genes of primary HIV clade C strains (termed SHIVenvC's); isolate, characterize, and sequence env genes from SHIVenvC-infected monkeys during disease progression, and perform diagnostic PCR to assess viral loads. The Specific Aims are: 1. To insert env genes derived from Zambian infants with rapidly progressive HIV clade C disease into multiply engineered SIVmac239-derived backbones to increase the replicative capacity of the chimeric SHIVenvC strains and to accelerate disease progression in primates. 2. To construct a fully heterologous SHIV based upon an SIV backbone that differs from SIVmac239, such as SIVsmE543-3, and by inserting an R5 HIV clade C env gene that differs from those used to construct the current SHIVenvC isolates. 3. To generate and characterize HIV clade C env mutants and to work closely with Project 1 to assess the molecular evolution of env in SHIVenvC-infected rhesus monkeys during disease progression. 4. To ensure that all experimental monkeys are free of other retrovirus infections prior to enrollment. A highly sensitive DMA PCR assay will be used to monitor for simian retrovirus type D (SRV/D) or simian Ivmphotropic virus type 1 (STLV-1). 5. To monitor all experimental animals in Core C by real-time RT-PCR and DNA PCR for plasma viral RNA or proviral DMA loads, respectively.

核心A的总体目标是通过以下方式为计划项目提供分子生物学专业知识和支持与所有三个项目和核心密切互动。核心A将构建新的类人猿-人类免疫缺陷病毒（SHIV）株编码原代HIV进化枝C株的R5 env基因（称为 SHIVenvC的）;分离，表征和测序来自SHIVenvC感染的猴的env基因，疾病进展，并进行诊断PCR以评估病毒载量。具体目标是： 1.将来自患有快速进展的HIV C分支疾病的赞比亚婴儿的env基因插入多重工程化的SIVmac 239衍生的主链，以增加SIVmac 239的复制能力。嵌合SHIVenvC菌株并加速灵长类动物中的疾病进展。 2.为了构建基于不同于SIVmac 239的SIV骨架的完全异源SHIV，例如SIVsmE 543 -3，并通过插入与用于构建当前SHIVenvC分离株。 3.产生和表征HIV进化枝C env突变体，并与项目1密切合作，评估 SHIVenvC感染恒河猴疾病进展过程中env的分子进化。 4.确保所有实验猴在入组前无其他逆转录病毒感染。一高灵敏度DMA PCR检测将用于监测猿猴逆转录病毒D型（SRV/D）或猴嗜光病毒1型（STLV-1）。 5.通过实时RT-PCR和DNA PCR监测核心C中所有实验动物的血浆病毒 RNA或前病毒DNA加载。