采用PCR技术扩增6个地理种群桑天牛的线粒体细胞色素氧化酶Ⅰ（mt DNA COⅠ）基因序列,基于mt DNA COⅠ基因序列变异探讨不同地理分布桑天牛种群间的遗传距离、系统发育以及遗传分化程度。对6个地理种群的33个桑天牛样本的mt DNA COⅠ基因扩增产物的测序结果表明共有19个位点发生变异,占序列总长的4.0%,其中来自河南省济源的桑天牛有12个变异位点,来自浙江省的桑天牛有9个变异位点,来自河北省4个地区的桑天牛的变异位点较少;河北省的4个地理种群的遗传距离较小,在0.001 3-0.002 6之间,河北、河南、浙江种群间的遗传距离较大,其中河南种群与河北种群的遗传距离又远远大于浙江种群与河北种群的遗传距离,后2大地理种群分布区域的海拔更接近。利用MEGA4.1软件构建的NJ系统进化树显示,6个地理种群形成河北、河南和浙江3大地理分布格局;从遗传分化程度上来看,桑天牛种群的3大地理分布格局不存在共享的单元型,遗传分化系数（FST）在0.804 1-0.892 2范围内,遗传分化程度较高。综合分析认为,不同地理种群桑天牛存在分化的原因除地理分布外,更重要的是生态条件的差异。
建立一整套适用于研究桑天牛线粒体细胞色素氧化酶I(mtDNA CO I)基因的PCR反应体系,研究不同地理分布的桑天牛遗传多样性。通过L_(16)(4~5)正交组合试验和单因素梯度试验对缓冲液(Mg^(2+))、dNTP、随机引物、TaqDNA聚合酶、模板DNA的浓度和退火温度、循环次数等影响PCR扩增的重要因素进行优化。试验结果表明,50μL反应体系及反应程序中各因素优化组合为:缓冲液(含Mg^(2+))0.9mmol/L,dNTP 0.15mmol/L,随机引物0.44μnol/L,TaqDNA聚合酶3.0U,模板DNA75ng;退火温度47℃,循环次数35次。
The effect of culture media,temperature and humidity on the mycelium growth,spores germination of Metarhizium MS01 strain,a high virulent strain from the soil and the pathogenicity against Anoplophora glabripennis larvae was tested. The result showed that the optimum culture medium was PPDA,the optimum temperature for the mycelium growth and spore germination was 26 ℃,and A. glabripennis larvae had the highest mortality from the fungus at this temperature. The optimum RH for the mycelium growth ranged from 95%to 100%. The speed and rate of spore germination increased with increasing of RH. The larvae died from the fugus most quickly and the mortality reached the highest under 100% RH.
