Xin Zhanguo, Wang Ming Li, Barkley Noelle A, Burow Gloria, Franks Cleve, Pederson Gary, Burke John
Plant Stress and Germplasm Development Unit, USDA-ARS, 3810 4th Street, Lubbock, TX 79415, USA.
BMC Plant Biol. 2008 Oct 14;8:103. doi: 10.1186/1471-2229-8-103.
Sorghum [Sorghum bicolor (L.) Moench] is ranked as the fifth most important grain crop and serves as a major food staple and fodder resource for much of the world, especially in arid and semi-arid regions. The recent surge in sorghum research is driven by its tolerance to drought/heat stresses and its strong potential as a bioenergy feedstock. Completion of the sorghum genome sequence has opened new avenues for sorghum functional genomics. However, the availability of genetic resources, specifically mutant lines, is limited. Chemical mutagenesis of sorghum germplasm, followed by screening for mutants altered in important agronomic traits, represents a rapid and effective means of addressing this limitation. Induced mutations in novel genes of interest can be efficiently assessed using the technique known as Targeting Induced Local Lesion IN Genomes (TILLING).
A sorghum mutant population consisting of 1,600 lines was generated from the inbred line BTx623 by treatment with the chemical agent ethyl methanesulfonate (EMS). Numerous phenotypes with altered morphological and agronomic traits were observed from M2 and M3 lines in the field. A subset of 768 mutant lines was analyzed by TILLING using four target genes. A total of five mutations were identified resulting in a calculated mutation density of 1/526 kb. Two of the mutations identified by TILLING and verified by sequencing were detected in the gene encoding caffeic acid O-methyltransferase (COMT) in two independent mutant lines. The two mutant lines segregated for the expected brown midrib (bmr) phenotype, a trait associated with altered lignin content and increased digestibility.
TILLING as a reverse genetic approach has been successfully applied to sorghum. The diversity of the mutant phenotypes observed in the field, and the density of induced mutations calculated from TILLING indicate that this mutant population represents a useful resource for members of the sorghum research community. Moreover, TILLING has been demonstrated to be applicable for sorghum functional genomics by evaluating a small subset of the EMS-induced mutant lines.
高粱[Sorghum bicolor (L.) Moench]是世界上第五大重要粮食作物，是世界许多地区，特别是干旱和半干旱地区的主要主食和饲料资源。近期高粱研究的激增是由其对干旱/热胁迫的耐受性以及作为生物能源原料的巨大潜力所驱动。高粱基因组序列的完成开辟了高粱功能基因组学的新途径。然而，遗传资源，特别是突变系的可用性有限。对高粱种质进行化学诱变，随后筛选重要农艺性状发生改变的突变体，是解决这一限制的快速有效方法。利用称为基因组靶向诱导局部损伤（TILLING）的技术，可以有效地评估新的感兴趣基因中的诱导突变。
通过用化学试剂甲磺酸乙酯（EMS）处理自交系BTx623，产生了一个由1600个品系组成的高粱突变群体。在田间观察到许多M2和M3品系的形态和农艺性状发生改变的表型。使用四个靶基因通过TILLING分析了768个突变体系的一个子集。总共鉴定出五个突变，计算得出的突变密度为1/526 kb。通过TILLING鉴定并经测序验证的两个突变在两个独立的突变系中存在于编码咖啡酸O-甲基转移酶（COMT）的基因中。这两个突变系分离出预期的棕色中脉（bmr）表型，该性状与木质素含量改变和消化率提高有关。
TILLING作为一种反向遗传学方法已成功应用于高粱。在田间观察到的突变体表型的多样性，以及从TILLING计算得出的诱导突变密度表明，这个突变群体是高粱研究界成员的有用资源。此外，通过评估一小部分EMS诱导的突变系，已证明TILLING适用于高粱功能基因组学。
