The establishment and application of positional cloning method for unknown gene in fish species of tetraploid origin.

四倍体鱼类未知基因定位克隆方法的建立及应用

• 批准号: 23248030
• 负责人: OKAMOTO Nobuaki
• 金额: $ 16.22万
• 依托单位: Tokyo University of Marine Science and Technology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 2011
• 资助国家: 日本
• 起止时间: 2011-11-18 至 2014-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-23248030/
• 关键词: ニジマス; 優性アルビノ; 候補遺伝子; 4倍性; 染色体歩行; ポジショナルクローニング; シンテニー解析; BACクローン; 4倍体起源種

As a first step towards the positional cloning of the gene responsible for autosomal dominant oculocutaneous albinism (OCA) in rainbow trout, we performed linkage analysis of SSLP147 markers associated with the dominant albino locus by using 1004 samples from analyzed family. And then, we found that the genetic distance between SSLP147 marker and the dominant albino locus was only 0.2 cM. As a second step, we started chromosome walking method using rainbow trout BAC libraries. As a results, we detected some BAC clones that have no recombinant between phenotypes and genotypes within 1004 samples. We determined whole sequences of the BAC clones and searched the candidate gene in the sequence. Unfortunately, we could not find the candidate gene of OCA. By candidate gene approach method, we isolated rainbow trout c-kit like gene that related with the pigment pattern mutant in zebrafish. We found some base pair substitutions in transcriptional regulatory region, but not in coding region.

作为定位克隆虹鳟显性眼皮肤白化病（OCA）基因的第一步，我们利用1004个家系样本对与显性白化病基因相关的SSLP 147标记进行了连锁分析。在此基础上，我们发现SSLP 147标记与显性白化病位点的遗传距离仅为0.2cM。作为第二步，我们开始使用虹鳟鱼BAC文库的染色体步移方法。结果，我们在1004份样本中检测到一些表型和基因型之间没有重组的BAC克隆。我们测定了BAC克隆的全序列，并在序列中寻找候选基因。不幸的是，我们没有找到OCA的候选基因。利用候选基因方法，分离了与斑马鱼色素模式突变相关的虹鳟c-kit like基因。我们在转录调控区发现了一些碱基对替换，但在编码区没有发现。

项目成果

ニジマス優性アルビノ原因遺伝子同定の試み

尝试鉴定导致白化虹鳟鱼显性的基因

• 发表时间: 2013
• 作者: Miyanaga;Y.;Kobayashi;A. and A. Murakami;二見邦彦・輿水江里子・本島佳苗・山根允文・坂本崇・岡本信明
• 通讯作者: 二見邦彦・輿水江里子・本島佳苗・山根允文・坂本崇・岡本信明