TILLING,即Targeting Induced Local Lesions IN Genomes(定向诱导基因组局部突变技术),是由美国Fred Hutchinson癌症研究中心Steven Henikoff领导的研究小组发展起来的,是一种全新的反向遗传学研究方法。TILLING技术借助高通量的检测手段,快速有效地从由化学诱变剂(EMS)诱变过的突变群体中鉴定出点突变。目前,TILLING已被应用于多种生物的研究中。本文系统介绍了TILLING技术的基本原理、技术路线及其技术优势,同时列举了TILLING的应用实例,并展望了TILLING技术的应用前景。
[Objective] The study aimed to identify Alternaria Nees from some areas of China at molecular level by analyzing the rDNA ITS sequence. [ Method ] The DNA sequences coding for the 5.8S rDNA and the flanking internal transcribed spacers （ ITS1 and ITS2） were amplified by PCR with universal primers ITS4 and ITS5 and subsequently sequenced for 34 Alternaria isolates from different areas of China. [Result] Sequences analysis showed that 5.8S rDNA was 159 bp and no variation in tested 34 isolates. There had variables sites in ITS. The isolates that had same sequences as A. tenuissima or A. alternata all put up eurytopicity to area and host. The variables sites of the isolates showed the diver- sity of Alternaria in the hosts of Oleaceae, Rosaceae and Solanaceae. At the same time that ITS could not clearly separated the isolates was indicated. The results indicated that the phylogenetic relationship were not closely related to the geographical origin and hosts of these isolates. [ Conclusion] The sequence analysis of ITS region could provide theory basis for the identification of Alternaria Nees..
