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斑皮蠹属昆虫个体小,形态高度相似,仅依据外部形态难以鉴定。用线粒体细胞色素氧化酶亚基I(COI)基因的通用引物,分别以黑斑皮蠹(Trogoderma glabrum)、花斑皮蠹(Trogoderma variabile)的DNA为模板,进行PCR扩增、测序。比较二者COI的碱基序列,找出所有序列多态性位点及相应的限制性内切酶,最终选取限制性内切酶AflⅡ对二者进行酶切。结果表明,黑斑皮蠹的COI序列可被限制酶AflⅡ切开,形成535bp和347bp两部分,而花斑皮蠹COI片段中因不具有AflⅡ酶切位点而不能被切开,从而将二者成功区分,实现对黑斑皮蠹和花斑皮蠹的快速分子鉴定。
Bark mites insect small, highly similar in form, difficult to identify based solely on external morphology. The common primers of mitochondrial cytochrome oxidase subunit I (COI) gene were amplified by PCR and sequenced respectively using the DNA of Trogoderma glabrum and Trogoderma variabile as templates. The nucleotide sequences of the two COIs were compared to find all the polymorphic sites and the corresponding restriction endonucleases. Finally, the restriction endonucleases Afl Ⅱ were selected to digest the two. The results showed that the COI sequence of B. mori was cleaved by the restriction enzyme AflII to form two parts of 535bp and 347bp. However, the COI fragment of P. dicotyledon could not be excised because of not having AflII cleavage site, Who successfully differentiated to achieve rapid molecular identification of Bemisia pallens and Pieridae.