Analysis of Marek's disease virus (MDV) genes responsible for oncogenes is by MDV and protection of chickens from Marek's disease.

对负责致癌基因的马立克氏病病毒 (MDV) 基因进行分析，通过 MDV 和保护鸡免受马立克氏病的侵害。

• 批准号: 03454187
• 负责人: HIRAI Kanji
• 金额: $ 4.1万
• 依托单位: TOKYO MEDICAL AND DENTAL UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1991
• 资助国家: 日本
• 起止时间: 1991 至 1992
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-03454187/
• 关键词: Marek's disease virus(MDV); Tumor; Sequence determination; PCR; Recombinant MDV; Restriction endonuclease map; Protein kinase; Vaccine; 遺伝子産物; マレック病ウイルス; cDNAクロ-ン; DNA塩基配列; 燐酸化蛋白; 糖蛋白; プロテインキナ-ゼ

(1).Sequence determination of the tumor-associated region of the Marek's disease virus type 1 (MDV1) genome and isolation of cDNA clones of transcripts from the region : (a)Twelve clones carrying the 132bp tandem direct repeat, tumor associated region, within the long inverted repeat region of MDV1 genome were isolated from a cDNA library constructed from chick embryo fibro-blasts infected with MDV1. Two open reading frames (ORF) were found in a cDNA clone encoding the mRNA from the 132bp repeat by sequence analysis. (b)Nucleotide sequence analysis of 9.3kb DNA of MDV1, including the tumor-associated 132bp repeat revealed the presence of 14 potential ORFs. Identification of the proteins encoded from these ORFs is under investigation.(2).Analysis of MDV1 genes related to vaccinal immunity: (a)The DNA sequence (8.9kb) covering about 70% of the short unique region (Us) and part of the short inverted repeat of MDV1-GA strain was determined.Out of nine potential ORFs within the 8.9kb sequen … More ce, four were found to be homologous to US10, US3, US2, and US6 (gD) of herpes simplex virus type 1. (b)The US3 was shown to encode MDV1 specific protein kinase by using a baculovirus vector(3).Construction of recombinant MDV1 expressing the lacZ gene of Escherichia coli: We attempted to insert the Escherichia coli lacZ gene at 22 sites into the MDV1 genome by homologous recombination. Among them, expression of the lacZ gene at four sites were found to be stable during serial passages in culture. Especially, the US10 and US3 genes were nonessential for induction of protective immunity as well as for in vitro viral growth.(4).Construction of the restriction endonuclease map of MDV2: A BamHI, EcoRI, and XhoI restriction endonuclease map of MDV2 DNA was constructed. We showed the presence of the collinear relationship among viral genomes of three serotypes.(5).Differentiation of oncogenic and nononcogenic strains of MDV1 by using PCR: Differentiation of oncogenic and nononcogenic strains of MDV1 was possible by PCR using the primers chosen from the sequence including the 132bp repeat. Less

（1）马立克氏病病毒1型（MDV 1）基因组肿瘤相关区的序列测定及转录本cDNA克隆的分离：（a）从MDV 1感染鸡胚成纤维细胞构建的cDNA文库中，分离到12个含有MDV 1基因组长反向重复序列（LRP）内132 bp的肿瘤相关区的cDNA克隆。序列分析表明，该基因的132 bp重复序列中含有两个开放阅读框（ORF）。（B）对MDV 1的9.3kb DNA（包括肿瘤相关的132 bp重复序列）进行核苷酸序列分析，发现存在14个潜在的ORF。这些ORF编码的蛋白质的鉴定正在研究中。（2）MDV 1-GA株与疫苗免疫相关基因的分析：（a）测定了MDV 1-GA株的8.9kb序列，该序列覆盖了MDV 1-GA株约70%的短独特区（short unique region，Us）和部分短反向重复序列，在该序列内的9个开放阅读框中，发现了9个与疫苗免疫相关的开放阅读框。 ...更多信息 ce，发现4个与单纯疱疹病毒1型的US 10、US 3、US 2和US 6（gD）同源。（B）通过使用杆状病毒载体显示US 3编码MDV 1特异性蛋白激酶（3）表达大肠杆菌lacZ基因的重组MDV 1的构建：我们尝试通过同源重组将大肠杆菌lacZ基因在22个位点处插入MDV 1基因组。其中，lacZ基因在四个位点的表达被发现是稳定的，在连续传代培养。特别是，US 10和US 3基因对于诱导保护性免疫以及对于体外病毒生长是非必需的。（4）、MDV 2限制性内切酶图谱的构建：构建了MDV 2 DNA的BamHI、EcoRI和XhoI限制性内切酶图谱。我们发现三种血清型的病毒基因组之间存在共线关系。（5）用PCR方法区分MDV 1的致癌株和非致癌株：用包含132 bp重复序列的引物进行PCR，可以区分MDV 1的致癌株和非致癌株。少

项目成果

Geng-Sheng zhu: "Differentiation of oncogenic and nononcogenic strains of Marek's disease virus type 1 by using polymerase chain reaction DNA amplification." AVIAN DISEASE. 36. 637-645 (1992)

Geng-Sheng Zhu：“通过聚合酶链式反应 DNA 扩增来区分 1 型马立克氏病病毒的致癌和非致癌株。”

Kazuhiro Nakajima: "Comparison of indirect immunofluorecence(IF) test with enzyme-linked immunosorbent assay(ELISA) in screening of hybridomas to very virulent Marek's disease virus." Microbiol. Immunol. Vol.36(2). 191-197 (1992)

Kazuhiro Nakajima：“间接免疫荧光 (IF) 测试与酶联免疫吸附测定 (ELISA) 在筛选杂交瘤与高毒力马立克氏病病毒方面的比较。”

Masashi Sakaguchi: "Sequence determination and genetic content of an8.9kb restriction fragment in the short unique region and the internal inverted repeat of Marek's disease virus." Virus Genes.

Masashi Sakaguchi：“马立克氏病病毒短独特区域和内部反向重复序列中 8.9kb 限制性片段的序列测定和遗传内容。”

Kazuhiro Nakajima: "Comparison of indirect immunofluorecence(IF)test with enzyme-linked immunosorbent assay(ELISA) in screening of hybridomas to very virulent Marek's disease virus." Microbiol.Immunol.36(2). 191-197 (1992)

Kazuhiro Nakajima：“间接免疫荧光 (IF) 测试与酶联免疫吸附测定 (ELISA) 在筛选杂交瘤与高毒力马立克氏病病毒方面的比较。”

Kazuhiro Nakajima: "Isolation of monoclonal antibodies against very virulent Marek's diesase virus." Microbiology and Immunilogy.

Kazuhiro Nakajima：“分离出针对剧毒马立克氏病病毒的单克隆抗体。”