A candidate-gene approach to clone the sorghum Brown midrib gene encoding caffeic acid O-methyltransferase
A candidate-gene approach to clone the sorghum Brown midrib gene encoding caffeic acid O-methyltransferase
复制标题
DOI：
10.1007/s00438-003-0824-4
复制
发表时间：
2003-05-01
影响因子：
3.1
通讯作者：
Vermerris, W
中科院分区：
文献类型：
--
作者：
Bout, S;Vermerris, W
由于木质素成分的变化，高粱的棕色中脉（bmr）突变体在叶子和茎中具有棕色维管组织。 bmr 突变体是通过硫酸二乙酯 (DES) 化学诱变产生的，类似于玉米的棕色中脉 (bin) 突变体。玉米和高粱棕色中脉突变体对于比较不同但进化相关的物种的木质素生物合成具有特别的价值。尽管高粱棕色中脉突变体于 1978 年首次被描述，但尚未克隆出任何棕色中脉基因。我们使用候选基因方法从高粱中克隆了第一个棕色中脉基因。基于对等位基因突变体 bmr12、bmr18 和 bmr26 的化学分析，我们假设这些突变体的木质素生物合成酶咖啡酸 O-甲基转移酶 (COMT) 的活性降低。 Northern 分析显示 COMT 基因的表达大幅降低。使用 PCR 从突变体和相应的野生型中克隆该基因。在所有三个突变体中，均鉴定出导致过早终止密码子的点突变：bmr12、bmr18 和 bmr26 因此是编码 COMT 的基因的突变等位基因。 RT-PCR表明所有三个突变体均表达突变等位基因，但相对于野生型对照水平低得多。为三个突变等位基因中的每一个开发了分子标记，以促进这些突变等位基因在遗传研究和育种计划中的使用。
The brown midrib (bmr) mutants of sorghum have brown vascular tissue in the leaves and stem as a result of changes in lignin composition. The bmr mutants were generated via chemical mutagenesis with diethyl sulfate (DES) and resemble the brown midrib (bin) mutants of maize. The maize and sorghum brown midrib mutants are of particular value for the comparison of lignin biosynthesis across different, yet evolutionarily related.. species. Although the sorghum brown midrib mutants were first described in 1978, none of the Brown midrib genes have been cloned. We have used a candidate-gene approach to clone the first Brown midrib gene from sorghum. Based on chemical analyses of the allelic mutants bmr12, bmr18 and bmr26, we hypothesized that these mutants had reduced activity of the lignin biosynthetic enzyme caffeic acid O-methyltransferase (COMT). After a northern analysis revealed strongly reduced expression of the COMT gene. the gene was cloned from the mutants and the corresponding wild types using PCR. In all three mutants,, point mutations resulting in premature stop codons were identified: bmr12, bmr18 and bmr26 are therefore mutant alleles of the gene encoding COMT. RT-PCR indicated that all three mutants express the mutant allele, but at much lower levels relative to the wild-type controls. Molecular markers were developed for each of the three mutant alleles to facilitate the use of these mutant alleles in genetic studies and breeding programs.