中国人兽共患病学报
中國人獸共患病學報
중국인수공환병학보
CHINESE JOURNAL OF ZOONOSES
2009年
12期
1181-1185
,共5页
小管福寿螺%肺囊%PCR%DNA%提纯%效果
小管福壽螺%肺囊%PCR%DNA%提純%效果
소관복수라%폐낭%PCR%DNA%제순%효과
Pomacea canaliculata%lung%PCR%DNA%extraction%efficiency
目的 以不同方法提纯小管福寿螺DNA, 用PCR检测以比较其效果.方法 将80只野外采集的小管福寿螺分8组,取出肺囊并剪碎后分别用QIAGEN、天根、OMEGA的基因组DNA抽提试剂盒,及异硫酸氰胍抽提法、树脂抽提法和树脂-二氧化硅抽提法提纯小管福寿螺基因组DNA,用PCR扩增福寿螺16s rDNA,测定目的 条带相对浓度以评价各方法的抽提效果.结果 各种方法提纯的基因组DNA均能用PCR扩增方法扩增出目的 条带,但目的 条带浓度有一定差异.其中,QIAGEN公司和OMEGA公司的抽提试剂盒、树脂抽提法和树脂-二氧化硅抽提法提纯的模板PCR扩增量较高,与其余4种抽提方法比较有统计学意义.试剂盒抽提法的单价成本是手工抽提法的8.4～57.1倍.结论 QIAGEN和OMEGA公司的基因组DNA抽提试剂盒的提纯效果好,但成本较高,适用于少量样本的抽提.树脂抽提法和树脂-二氧化硅抽提法从提纯效果、成本等方面评价,较适用于大量样本抽提处理工作.异硫酸氰胍抽提法,抽提快,成本低但所用试剂对身体有毒害,较适用于大量样本或应急样本的处理.
目的 以不同方法提純小管福壽螺DNA, 用PCR檢測以比較其效果.方法 將80隻野外採集的小管福壽螺分8組,取齣肺囊併剪碎後分彆用QIAGEN、天根、OMEGA的基因組DNA抽提試劑盒,及異硫痠氰胍抽提法、樹脂抽提法和樹脂-二氧化硅抽提法提純小管福壽螺基因組DNA,用PCR擴增福壽螺16s rDNA,測定目的 條帶相對濃度以評價各方法的抽提效果.結果 各種方法提純的基因組DNA均能用PCR擴增方法擴增齣目的 條帶,但目的 條帶濃度有一定差異.其中,QIAGEN公司和OMEGA公司的抽提試劑盒、樹脂抽提法和樹脂-二氧化硅抽提法提純的模闆PCR擴增量較高,與其餘4種抽提方法比較有統計學意義.試劑盒抽提法的單價成本是手工抽提法的8.4～57.1倍.結論 QIAGEN和OMEGA公司的基因組DNA抽提試劑盒的提純效果好,但成本較高,適用于少量樣本的抽提.樹脂抽提法和樹脂-二氧化硅抽提法從提純效果、成本等方麵評價,較適用于大量樣本抽提處理工作.異硫痠氰胍抽提法,抽提快,成本低但所用試劑對身體有毒害,較適用于大量樣本或應急樣本的處理.
목적 이불동방법제순소관복수라DNA, 용PCR검측이비교기효과.방법 장80지야외채집적소관복수라분8조,취출폐낭병전쇄후분별용QIAGEN、천근、OMEGA적기인조DNA추제시제합,급이류산청고추제법、수지추제법화수지-이양화규추제법제순소관복수라기인조DNA,용PCR확증복수라16s rDNA,측정목적 조대상대농도이평개각방법적추제효과.결과 각충방법제순적기인조DNA균능용PCR확증방법확증출목적 조대,단목적 조대농도유일정차이.기중,QIAGEN공사화OMEGA공사적추제시제합、수지추제법화수지-이양화규추제법제순적모판PCR확증량교고,여기여4충추제방법비교유통계학의의.시제합추제법적단개성본시수공추제법적8.4～57.1배.결론 QIAGEN화OMEGA공사적기인조DNA추제시제합적제순효과호,단성본교고,괄용우소량양본적추제.수지추제법화수지-이양화규추제법종제순효과、성본등방면평개,교괄용우대량양본추제처리공작.이류산청고추제법,추제쾌,성본저단소용시제대신체유독해,교괄용우대량양본혹응급양본적처리.
To compare the efficiency of methods of DNA extraction from lungs of Pomacea canaliculataused in PCR assay, 80 P.canaliculata collected in field were divided into 8 groups and the lungs of each snail were separated from the soft body. Six methods of DNA extraction from lungs of P. canaliculata were used to extract DNA from lungs, i.e. With Qiagen, Tiangen,and Omega commercial DNA extraction kits, guanidine thiocyanate method, Chelex 100 resin method and Chelex-silica particle method. The 16S rDNA of C.canaliculata was amplified by PCR and the concentration of PCR-products relative to marker was determined in order to evaluate the efficiency of each method. It was demonstrated that each method was valid to extract DNA from lungs used in PCR assay, but the concentrations of PCR-products were different. The concentrations of PCR-products obtained by Qiangen kit, Omega kit, Chelex 100 resin method and Chelex-silica particle method were significantly higher than those of other 4 methods of DNA extraction, in which Qiangen and Omega kits were suitable for small sample size. In term of efficiency and cost, Chelex 100 method and Chelex-silica particle method were feasible for large sample scale, while the guanidine thiocyanate method was preferred due to its fast extraction and low cost, but on account of its toxicity, it is used in urgent status or in large scale of sample extraction.
