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Abstract
Black Queen Cell Virus (BQCV) is one of the most prevalent viruses in honeybee, and could not be recognized by any infected symptoms in the adult bees. Thus, a newly developed method for rapid RNA isolation and cDNA generation from honeybee samples will be useful and easy to evaluate the presence of BQCV. A significant improvement by only one-step RNA isolation for 10 minutes made possible to detect BQCV RNAs at high quantity. cDNAs were also generated directly from isolated BQCV RNAs by different primer sets for 1 minutes and were subsequently applied to Ultra-Rapid Real-Time PCR by using microchip of 6μl reaction volume with extremely short time in each step of PCR. This system provided the ultra-high speed reaction (30 cycles in less than 10 min) including melting temperature analysis for amplified BQCV products. These results suggest that BQCV detection can be completed within 6 min 42 sec, and the Ultra-Rapid Real-time PCR is sensitive, reliable and time-saving for monitoring BQCV in honeybee.
Published Version
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