豆象干標本基因組DNA提取方法的改進

李金慶 魯閩 粟智平 段效輝 王穎 孫超 盧經

摘要[目的] 篩選豆象干標本基因組DNA的有效提取方法。[方法] 以口岸截獲時間不等的豆象干標本為材料，在傳統方法上加以改進，對從干標本中提取豆象基因組DNA的方法進行了研究。將其所提取的基因組DNA的純度和濃度與CTAB法和SDS法進行了比較，并對DNA進行了完整性及PCR驗證。[結果] 與CTAB法和SDS法相比，豆象干標本提取方法提取的DNA OD260/OD280值均在1.7～1.9，且濃度合適。DNA完整性及PCR驗證結果表明，豆象干標本提取方法比較適合后續PCR試驗的開展。[結論] 豆象干標本提取方法適合該研究中豆象干標本基因組DNA的提取，可推廣到檢驗檢疫工作中截獲豆象的檢疫鑒定中，以縮短檢驗檢疫工作周期，提高檢驗檢疫工作準確性。

關鍵詞豆象干標本；DNA提取；基因組DNA

中圖分類號S412文獻標識碼A文章編號0517-6611（2017）14-0123-03

Abstract[Objective] To select the effective DNA extraction method for dried samples of bean weevil insects.[Method] The method for extracting genome DNA was studied on the basis of traditional methods by using dried bean weevil insects of different intercepted times as materials.The purity and concentration of the extracted genomic DNA samples were detected and compared with CTAB and SDS method， then the extracted genomic DNA samples were further verified by the DNA integrity and PCR identification.[Result] Compared with CTAB and SDS method， the extracted genomic DNA samples had appropriate OD260/OD280 values between 1.7-1.9 and appropriate concentrations. The DNA integrity and PCR identification results showed that the genomic DNA extraction method for dried bean weevil samples was more suitable for subsequent PCR experiments. [Conclusion] Dried bean weevil DNA extration method is suitable for extrating bean weevil insectsgenomic DNA. This method can be extended to the quarantine and identification of intercepted bean weevil samples in CIQ work， in order to shorten the period and improve the accuracy of CIQ work.

Key wordsDried bean weevil sample；DNA extraction；Genomic DNA

近年來，分子生物學技術被大量應用于檢驗檢疫工作中檢疫性昆蟲的鑒定等[1-5]，這些分子生物學鑒定技術快速有效，可有效彌補昆蟲形態學鑒定中早期形態（包括卵、幼蟲/若蟲、蛹）與殘缺蟲體難以辨認、鑒定結果受主觀影響大、鑒定周期較長等缺點[6]。進行分子生物學鑒定時，昆蟲中有效基因組DNA的提取是鑒定成功與否的前提[7]。在以往的研究中，大多數研究者所用的昆蟲基因組DNA 提取材料基本上是新鮮樣品或超低溫冷凍標本。但是在實際工作中，針對稀有類群或檢疫對象經常面臨難以得到新鮮材料的問題，往往需要利用標本館或從其他渠道得到的干標本提取基因組DNA。……