In situ analysis of latency-associated transcripts as an indicator of latent infection of Marek's disease virus.

潜伏相关转录本的原位分析作为马立克氏病病毒潜伏感染的指标。

基本信息

批准号：
08660354
负责人：
ENDOH Daiji
金额：
$ 1.54万
依托单位：
HOKKAIDO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08660354/
关键词：
Marek's disease virus latent infection in situ hybridization digoxienin-labeled probe RT-PCR hybrid histochemistry latency-associated transcript MDV-transformed cell line in situハイブリダイゼーション

项目摘要

The purpose of this study is the transcriptional analysis of latently MDV-infected cells. In 1996 fiscal year, we made modification and became skillful in the methods for the analysis. Since low copies of MDV-trancripts would express in latently infected cells, we estimated reverse transcriptase-polymerase chain reaction (RT-PCR) , in situ hybridization and immunohistochemical techniques. In 1997 fiscal year, we detected newly identified latency-associated transcript (LAT) from MDV-transformed cells, which is known as a model of latent infection of MDV.In the MDV-transformed cells, spontaneously activated MDV is rarely included while it may influence the result of transcriptional analysis. We showed that a population consisting of 10^3 MDV-transformed cells probably did not include spontaneously activated MDV from immunohistochemical and RT-PCR-limiting-dilution analysis of MDV-transformed cells. In investigated genes (ICP4, ICP27, po1, TK,US3, A41, gA,gB,UL50, VP16andSORF2) a transcript corresponding to the ICP4 sequence was detected as a 0.7kbp RT-PCR product in 10^3 cells of MSB-1, RPL-1, HPRS-1, MOGA-1, MOGA-2, MSB-1 and MTB-1 but not in the retrovirus-transformed 1104B1 cell line. The transcript corresponding to the 0.7 kbp RT-PCR product suggested to be a common transcript in latently infected MDV-transformed cells, while most of the genes did not transcribe in these cells.

这项研究的目的是对潜在的MDV感染细胞的转录分析。在1996财政年度，我们进行了修改，并在分析方法上变得熟练。由于MDV-Trancripts的低拷贝将在潜在感染的细胞中表达，因此我们估计了逆转录酶 - 聚合酶链反应（RT-PCR），原位杂交和免疫组织化学技术。在1997财政年度，我们从MDV转化的细胞中检测到新发现的与潜伏相关的转录（LAT），该细胞被称为MDV转化细胞的潜在感染模型，自发激活的MDV很少被包括在内，但可能会影响转录分析的结果。我们表明，由10^3 MDV转换细胞组成的人群可能不包括对MDV转换细胞的免疫组织化学和RT-PCR限速分析的自发激活的MDV。在研究的基因（ICP4，ICP27，PO1，TK，US3，A41，GA，GB，GB，UL50，VP16和SORF2）中，在10 0.7kbp RT-PCR产物中检测到与ICP4序列相对应的转录本，在10^3个细胞中，MSB-1，RPLS-1，RPLS-1，HPRS-1，MOGA-1，MOGA-1，MOGA，MSB，MSM MSB，MSB-1逆转录病毒转换的1104B1细胞系。与0.7 kbp RT-PCR产物相对应的转录本建议是在潜在受感染的MDV转换细胞中的常见转录本，而大多数基因在这些细胞中没有转录。