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Cellular and Molecular Breeding of Sesame.

芝麻的细胞和分子育种。

基本信息

批准号：
09660003
负责人：
YAMADA Kyoji
金额：
$ 2.05万
依托单位：
Toyama University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

(i) We isolated full-length cDNA clones coding for two fatty acid desaturases from sesame, Sesamum indicum : a microsomal omega-6 desaturase which is required for conversion of oleic to linoleic acids and a plastid-localiz- ed omega-3 desaturase which is required for conversion of linoleic to linolenic acids. Sequence analysis indicated that the former cDNA clone included an open reading frame (ORF) of 383 amino acids, while the latter clone that of 447 amino acids. We also cloned a cDNA which encodes about one-third of a plasatid-localized omega-6 fatty acid desaturase.(ii) Northern blot analyses showed changes in the transcription level of the above three fatty acid desaturases at various developmental stages and among different organs in sesame. A gene for an microsomal omega-6 desaturase was found to be transcribed at high level specifically in seeds at their immature stages, whereas the transcripts of plastid-localized type of both omega-6 and omega-3 desaturases were almost undetectable in various organs at all developmental stages.(iii) We have succeeded in cloning a genomic DNA of an microsomal omega-6 fatty acid desaturase of sesame, determined its nucleotide sequences, and then found that a 1.6 kbp intron was located at only 25 bp before the initiation codon of the coding region. At least 14 cis-regulatory elements, such as E-box, which are specific sequence motifs known to be effective for high expression of specific genes during seed development, were detected within the intron.(iv) We tried to transform sesame plants by infection of Agrobacterium. when infected with A.tumefaciens containing the MAT vector (pMAT21), seedlings of a wild species, Sesamum schinzianum frequently produced crown galls. Incubation of the calli derived from the goals resulted in regenera- tion of plantlets at high frequency. Confirmation of transformation in such regenerated plants is now in progress.

(i)我们分离全长cDNA克隆编码的两个脂肪酸去饱和酶从芝麻，芝麻：一个微粒体ω-6去饱和酶，这是所需的油酸转化为亚油酸和质体定位的艾德ω-3去饱和酶，这是所需的亚油酸转化为亚麻酸。序列分析表明，前者含有383个氨基酸的开放阅读框架，后者含有447个氨基酸的开放阅读框架。我们还克隆了一个cDNA，它编码约三分之一的质体定位的ω-6脂肪酸去饱和酶。(ii)北方杂交分析表明，上述三种脂肪酸脱氢酶在芝麻不同发育时期和不同器官间的转录水平存在变化。发现微粒体ω-6去饱和酶的基因在未成熟阶段的种子中特异性地以高水平转录，而质体定位型的ω-6和ω-3去饱和酶的转录物在所有发育阶段的各种器官中几乎检测不到。(iii)我们已经成功地克隆了芝麻微粒体ω-6脂肪酸去饱和酶的基因组DNA，测定了其核苷酸序列，然后发现一个1.6 kbp的内含子位于编码区的起始密码子前仅25 bp处。在内含子中检测到至少14个顺式调控元件，如E-box，它们是已知在种子发育期间对特定基因的高表达有效的特定序列基序。(iv)我们尝试通过农杆菌感染来转化芝麻植株。当用含有MAT载体（pMAT 21）的根癌农杆菌感染时，野生种Sesamumschinzianum的幼苗经常产生冠瘿。对来自目标的愈伤组织的孵育导致高频率的植株再生。目前正在确认这种再生植物的转化。