Wiersma P A, Schmiemann M G, Condie J A, Crosby W L, Moloney M M
Department of Biological Sciences, University of Calgary, Canada.
Mol Gen Genet. 1989 Nov;219(3):413-20. doi: 10.1007/BF00259614.
An acetolactate synthase gene was isolated and characterized from Brassica napus. This B. napus acetolactate synthase gene encodes a deduced polypeptide sequence of 637 amino acids which is 85% homologous to the corresponding proposed gene product from Arabidopsis thaliana. Peptide domains recently associated with herbicide resistance/sensitivity are conserved between the two sequences. From Southern analysis we conclude that the gene isolated is one member of a multigene acetolactate synthase gene family comprising four or five members. A probe spanning the presumptive transit peptide sequence of this gene was shown by Southern analysis to hybridize to a unique sequence in the B. napus genome. This unique probe was used to analyse DNA from B. campestris and B. oleracea, the presumed progenitors of B. napus. On the basis of restriction fragment length polymorphism, we conclude that the B. napus gene isolated here originated in B. campestris. Total acetolactate synthase-homologous transcripts were analysed in a variety of B. napus tissues, and showed preferential accumulation in rapidly growing material. The genomic clone was mutated in vitro at codon 173 to replace a proline residue with serine. This was re-introduced into plants, using Agrobacterium vectors, producing a herbicide-resistant phenotype which is characteristic of the predicted gene product.
从甘蓝型油菜中分离并鉴定了一个乙酰乳酸合酶基因。该甘蓝型油菜乙酰乳酸合酶基因编码一个由637个氨基酸组成的推导多肽序列，与拟南芥相应的推测基因产物具有85%的同源性。最近与除草剂抗性/敏感性相关的肽结构域在这两个序列之间是保守的。通过Southern分析，我们得出所分离的基因是一个由四到五个成员组成的多基因乙酰乳酸合酶基因家族的一员。Southern分析表明，一个跨越该基因推定转运肽序列的探针与甘蓝型油菜基因组中的一个独特序列杂交。这个独特的探针被用来分析油菜的假定祖先——白菜和甘蓝的DNA。基于限制性片段长度多态性，我们得出这里分离的甘蓝型油菜基因起源于白菜。在多种甘蓝型油菜组织中分析了总乙酰乳酸合酶同源转录本，结果表明其在快速生长的材料中优先积累。在体外将基因组克隆的第173位密码子的脯氨酸残基突变为丝氨酸。利用农杆菌载体将其重新导入植物中，产生了一种除草剂抗性表型，这是预测基因产物的特征。
