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Applied and Environmental Microbiology, August 1999, p. 3714-3716, Vol. 65, No. 8
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Multiplex PCR Screening To Detect cry9 Genes in Bacillus thuringiensis Strains

Eitan Ben-Dov,¹ Qingfeng Wang,¹ Arieh Zaritsky,¹ Robert Manasherob,¹ Ze'ev Barak,¹ Bert Schneider,¹ Aloviddin Khamraev,² Mukhtar Baizhanov,³ Victor Glupov,⁴ and Yoel Margalith^1,*

Department of Life Sciences, Ben-Gurion University of the Negev, Be'er-Sheva 84105, Israel¹; Institute of Zoology, Uzbek Academy of Science, Tashkent 700095, Uzbekistan²; Institute of Zoology, Kazakhstan Academy of Science, Almaty 480032, Kazakhstan³; and Institute of Systematic and Animal Ecology, Siberian Branch of Academic Science, Novosibirsk 6300091, Russia⁴

Received 3 March 1999/Accepted 18 May 1999

An extended PCR method was established to rapidly identify and classify Bacillus thuringiensis strains containing cry (crystal protein) genes toxic to lepidopteran, coleopteran, and dipteran pests (Ben-Dov et al., Appl. Environ. Microbiol. 63:4883-4890, 1997). To optimize identification of all reported cry genes, this methodology needs a complete PCR set of primers. In the study reported here, a set of universal (Un9) and specific primers for multiplex rapid screening for all four known genes from the cry9 group was designed. PCR analyses were performed for cry9 genes on 16 standard strains and 215 field isolates of B. thuringiensis. Among the standard strains, only B. thuringiensis subsp. aizawai HD-133, which harbors cry1 and cry2 genes, was positive with Un9 but negative to all four specific primers for cry9 genes. DNA of 22 field-collected isolates was also found to be positive with Un9. These isolates were classified into three cry9 profiles using specific primers; all of them harbor cry1 and cry2. This newly designed set of primers complements the existing PCR methodology for most currently known cry genes.

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^* Corresponding author. Mailing address: Department of Life Sciences, Ben-Gurion University of the Negev, P.O. Box 653, Be'er-Sheva 84105, Israel. Phone: 972-7-6461.340 or 972-7-6472.642. Fax: 972-7-6472.890. E-mail: yoelm{at}bgumail.bgu.ac.il.

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Applied and Environmental Microbiology, August 1999, p. 3714-3716, Vol. 65, No. 8
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

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