Costa Ana S M B, Thomas D John I, Eastwood Daniel, Cutler Simon B, Bailey Andy M, Foster Gary D, Mills Peter R, Challen Micheal P
Warwick HRI, University of Warwick, Wellesbourne, Warwickshire, CV35 9EF, U.K.
J Microbiol Biotechnol. 2009 Mar;19(3):271-6.
Functional gene studies in the cultivated white button mushroom Agaricus bisporus have been constrained by the absence of effective gene-silencing tools. Using two endogenous genes from A. bisporus, we have tested the utility of dsRNA hairpin constructs to mediate downregulation of specific genes. Hairpin constructs for genes encoding orotidine 5'- monophosphate decarboxylase (URA3) and carboxin resistance (CBX) were introduced into A. bisporus using Agrobacteriummediated transfection. Although predicted changes in phenotype were not observed in vitro, quantitative-PCR analyses indicated unambiguously that transcripts in several transformants were substantially reduced compared with the non-transformed controls. Interestingly, some hairpin transformants exhibited increased transcription of target genes. Our observations show that hairpin transgenic sequences can mediate downregulation of A. bisporus endogenous genes and that the technology has the potential to expedite functional genomics of the mushroom.
由于缺乏有效的基因沉默工具，双孢蘑菇（Agaricus bisporus）的功能基因研究受到了限制。我们利用双孢蘑菇的两个内源基因，测试了双链RNA发夹结构介导特定基因下调的效用。通过农杆菌介导的转染，将编码乳清苷5'-单磷酸脱羧酶（URA3）和羧菌灵抗性（CBX）的基因的发夹结构导入双孢蘑菇。尽管在体外未观察到预测的表型变化，但定量PCR分析明确表明，与未转化的对照相比，几个转化体中的转录本大幅减少。有趣的是，一些发夹转化体表现出靶基因转录增加。我们的观察结果表明，发夹转基因序列可以介导双孢蘑菇内源基因的下调，并且该技术有潜力加快蘑菇功能基因组学的研究。
