ANALYSIS OF DWARF GENE(UZ)OF STRAIN UZU-AKASHINNRIKI OF BARLEY

大麦UZU-AKASHINRIKI矮化基因(UZ)分析

基本信息

批准号：
63540534
负责人：
SAKURAI Naoki
金额：
$ 1.15万
依托单位：
HIROSHIMA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1988
资助国家：
日本
起止时间：
1988 至 1989
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-63540534/
关键词：
AUXIN BARLEY DWARF mRNA TRANSAMINASE RACEMASE 遺伝子オーキシントリプトファン C-DNA m-RNA

项目摘要

Normal barley strain (Nami Akashinnriki) and dwarf strain (uzu Akashinriki) were grown under the dark. The coleopitles were harvested on day 3, when the growth rate of the normal strain was ca. two times as fast as that dwarf strain. mRNA was extracted and purified from the coleoptile segments (1 cm in length, excised from 5 mm below the tip). mRNA thus obtained was introduced into the in vitro translation system using wheat germ extracts with radio labeled ^<35>S-methionine or ^3H-leucine. The transcripts were dissolved on two-dimensional gel electrophoresis. The comparison of the fluorogram of the normal strain with that of the dwarf strain revealed that there were 11 species of proteins distinctly appeared in the transcripts from mRNA of the nominal strain, but were not or little transcribed from mRNA of the dwarf strain.IAA was added to the in vitro translation system to see the effect of exogenously applied IAA on the translation process. IAA did not affect the translation in stra … More in uzu either using methionine or leucine, while in stmin Nami, IAA substantially inhibited the incorporation of labeled leucine into the proteins but not the incorporation of methilnine. The unexpected results remained to be answered.As reported previously, strain uzu produced less IAA than strain Nami. To see which step for IAA bioshynthesis was impeded in strain uzu, crude enzyme preparation was extracted from the coleoptile segments of uzu and Nami. IAA was produced from D-tryptophan in both strains. The production was inhibited by D-cycloserine (an inhibitor of D-transaminase), suggesting that IAA is produced through D-tryptophan in both strains. Next, racemase activity which converts L-trp to D-trp was assayed. pH optimum for the racemase of Nami was 7.6 while that of uzu was over 9.7. The difference of the optimum pH for the racemase between Nami and uzu strongly suggested that the amino acid composition of racemase of uzu was different from that of Nami, and DNA sequence for racemase of uzu was also different from that of Nami. The isolation of the racemase was now undertaken by ion- exchange and gel permeation chromatography. Purified enzyme and its amino acid composition will afford a great facility for the analysis of dwar gene of uzu. Less

正常的大麦菌株（Nami Akashinnriki）和矮人菌株（Uzu Akashinriki）在黑暗下生长。当正常应变的生长速率大约时，在第3天收获了小组。两倍是矮人菌株。提取mRNA并从鞘翅目段纯化（长度为1 cm，从尖端下方5 mm处出色）。这样获得的mRNA使用带有无线电标记 ^<35> s-methionine或 ^3H-亮氨酸的小麦胚芽提取物引入体外翻译系统中。转录本溶解在二维凝胶电泳上。正常应变与矮菌株的荧光图的比较表明，从名义菌株的mRNA中，有11种蛋白质明显出现在转录本中，但并未从矮人菌株的mRNA中转录或很少转录。IAA的mRNA被添加到体外翻译系统中，以查看在易于应用IAA的效应。 IAA并不影响稻草中的翻译……更多的是使用甲基氨酸或亮氨酸在Uzu中影响，而在Stmin Nami中，IAA中的IAA实质上抑制了掺入的工业标记的亮氨酸，而不是融合的工业方法。意外的结果仍有待回答。正如先前报道的那样，UZU产生的IAA少于NAMI菌株。为了查看在UZU菌株中阻碍了IAA生物合成的哪一步，从Uzu和Nami的鞘臂段中提取了粗酶制剂。 IAA都是由D- tryptophan在两种菌株中生产的。 D-胞克氨烯（D-Transami的抑制剂）抑制了产量，这表明IAA是通过两种菌株中D- tryptophan生产的。接下来，分配了将L-TRP转换为D-TRP的Racemase活动。 NAMI种族酶的最佳pH值为7.6，而UZU的pH值超过9.7。 NAMI和UZU之间的种族酶的最佳pH差异很强地表明，UZU的Racemase的氨基酸组成与NAMI的氨基酸组成不同，而UZU的Racemase的DNA序列也与NAMI的DNA序列不同。现在由离子交换和凝胶渗透色谱法进行了种族主义酶的分离。纯化的酶及其氨基酸组成将为分析Uzu的矮基因提供一个很好的设施。较少的