Basic approach for improvement of breeding by using chromosome vector

利用染色体载体改良育种的基本途径

• 批准号: 07556137
• 负责人: MAEKAWA Hideaki
• 金额: $ 3.01万
• 依托单位: NATIONAL INSTITUTE OF INFECTIOUS DESEASES
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07556137/
• 关键词: CHROMOSOME; FISH; transposable element; RAPD; RFLP; CENP-A; temperature sensitive mutant of baculovirus; BOMBYX MORI; レトロトランスポゾン; 温度感受性株; バキュロウイルス; テロメア; 培養細胞; 二価染色体; 動原体; 倍数体; 転座

Identification of markers for almost all 28 linkage groups of the silkworm, Bombyx mori, was determined by analysis of Random Amplified Polymorphic DNA (RAPD) and Restriction Fragment Length Polymorphism (RFLP) of sequence from cDNA libraries. Identification of specific chromosomes was carried out by using mutated bivalent chromosomes at the metaphase of the first meiosis in polyploid oocytes according to size and the procedure for preparation of those chromosomes was improved by using polytron. Probes of reported elements including retrotransposons located on the ends of chromosomes and mariner elements clustered on specific chromosomes were also used as references for identification by Fluorescence In Situ Hybridization (FISH). Additional characterization was attempted by an indirect fluorescence method using an anti-B.mori CENP-A-like protein antibody in order to detect centromeres which are considered to be diffuse. This antibody bound to centromeres of human chromosomes but to a different region in B.mori. A temperature sensitive mutant of the nuclear polyhedrosis virus of B.mori, a kind of baculovirus, was isolated as a candidate for a gene introduction vector, and combined with a retrotransposon, BMC1. This construct was shown to be suitable as an expression vector. A rapid characterization procedure for cultured cells was established by PCR amplification of three genes specific to B.mori.

通过分析来自cDNA文库的序列的随机扩增多态性DNA（RAPD）和限制性片段长度多态性（RFLP），确定了几乎所有28个蚕的连锁基团的标记。通过使用多倍体卵母细胞中第一个减数分裂的中期，根据大小，使用突变的二价染色体来鉴定特异性染色体，并通过使用多晶龙改善了这些染色体的准备程序。报告的元素的探针包括位于特定染色体上聚集在染色体和水手元素末端的逆转录子，也被用作通过荧光原位杂交（FISH）鉴定的参考文献。使用抗B.mori CENP-A样蛋白抗体的间接荧光方法尝试了其他表征，以检测被认为是弥漫的中心粒。这种抗体与人类染色体的centromeres结合，但在b.mori中的另一个区域。 B. -Mori的核多面病毒的温度敏感突变体是一种杆状病毒，分离为基因引入载体的候选者，并与逆转录转座子BMC1结合在一起。该构建体被证明是作为表达矢量的。通过PCR扩增特定于B. -Mori的PCR扩增，建立了培养细胞的快速表征程序。

项目成果

伴野豊: "Chromosomal features at different meiotic stages during spermatogenesis in Bombyx mori" J.Silkworms.Sericologia. 36(1). 51-57 (1996)

Yutaka Banno：“家蚕精子发生过程中不同减数分裂阶段的染色体特征”J.Silkworms.Sericologia 36(1) (1996)。

BANNO,Y.: "GENETICAL STUDIES ON THE SEX-LINKED NON-MOLTING MUTATION,NM-S,OF BOMBYX MORI." J.SERIC.SCI.JPN.66-5. 351-355 (1997)

BANNO,Y.：“家蚕性连锁非蜕皮突变 NM-S 的遗传学研究。”

Takahashi, H.: "A NEW FAMILY OF SITE-SPECIFIC RETROTRANSPOSONS.SART1,IS INSERTED INTO TELOMERIC REPEATS OF THE SILKWORM,BOMBYX MORI." Nicl.Acids Res.25. 1578-1584 (1997)

Takahashi, H.：“一个新的位点特异性逆转录转座子家族。SART1，被插入到桑蚕的端粒重复序列中。”

Fujiwara, H.: "DEVELOPMENTAL PROFILES OF WING IMAGINAL DISCS OF FLUGELLOS(FL),A WINGLESS MUTANT OF THE SILKWORM,BOMBYX MORI." Dev Genes Evol.207. 12-18 (1997)

Fujiwara, H.：“FLUGELLOS（FL）翅成虫盘的发育概况，蚕是家蚕的无翅突变体。”

FUJIWARA,H.: "DEVELOPMENTAL PROFILES OF WING IMAGINAL DISCS OF FLUGELLOS (FL), A WINGLESS MUTANT OF THE SILKWORM,BOMBYX MORI." DEV.GENES.EVOL.207. 12-18 (1997)

FUJIWARA,H.：“FLUGELLOS（佛罗里达州）翅成虫盘的发育概况，FLUGELLOS 是家蚕的无翅突变体。”