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solarbio-DNA病毒基因組提取試劑盒
本試劑(ji)(ji)盒適合于從血清(qing)、細胞上清(qing)、淋巴(ba)液(ye)(ye)中(zhong)(zhong)(zhong)提(ti)取(qu)(qu)DNA病毒(du)基(ji)因(yin)組，不適合于RNA病毒(du)基(ji)因(yin)組的(de)(de)提(ti)取(qu)(qu)。使(shi)用本試劑(ji)(ji)盒提(ti)取(qu)(qu)的(de)(de)基(ji)因(yin)組DNA可(ke)用于各(ge)種常規(gui)操作，包括(kuo)酶切(qie)、PCR、文庫(ku)構(gou)建、Southern雜交等實驗。操作步(bu)驟(zou)： 使(shi)用前請先在漂(piao)洗(xi)(xi)液(ye)(ye)中(zhong)(zhong)(zhong)加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)無(wu)(wu)水乙(yi)醇，加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)體積請參照瓶上的(de)(de)標簽(qian)。所有離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)步(bu)驟(zou)均(jun)為(wei)使(shi)用臺式離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)機在室(shi)溫(wen)下離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)。1、取(qu)(qu)病毒(du)上清(qing)液(ye)(ye)0.5ml，12000rpm離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)5min，盡量(liang)吸(xi)(xi)盡上清(qing)使(shi)用，棄去沉淀。2、向(xiang)病毒(du)上清(qing)中(zhong)(zhong)(zhong)加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)20ul 10mg/ml的(de)(de)蛋白酶K，充分(fen)混(hun)勻(yun)，65℃消化10-20min，期間可(ke)顛倒離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)管(guan)混(hun)勻(yun)數(shu)次。3、向(xiang)管(guan)中(zhong)(zhong)(zhong)加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)500ul結合液(ye)(ye)，充分(fen)混(hun)勻(yun)。再向(xiang)管(guan)中(zhong)(zhong)(zhong)加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)400ul無(wu)(wu)水乙(yi)醇，充分(fen)混(hun)勻(yun)，此時(shi)可(ke)能會(hui)(hui)出現絮(xu)(xu)狀沉淀，不影響(xiang)DNA的(de)(de)提(ti)取(qu)(qu)，可(ke)將溶液(ye)(ye)和絮(xu)(xu)狀沉淀都加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)，靜(jing)置(zhi)2min。（吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)的(de)(de)大(da)容積為(wei)750ul，可(ke)分(fen)兩次加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)。一(yi)次吸(xi)(xi)附(fu)(fu)(fu)完(wan)離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)后再將余(yu)下的(de)(de)混(hun)合液(ye)(ye)體加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)靜(jing)置(zhi)離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)。） 4、 12000rpm離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)2min，棄廢液(ye)(ye)，將吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)入(ru)(ru)(ru)(ru)(ru)收(shou)集(ji)管(guan)中(zhong)(zhong)(zhong)。5、向(xiang)吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)700ul漂(piao)洗(xi)(xi)液(ye)(ye)(使(shi)用前請先檢查是否已加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)無(wu)(wu)水乙(yi)醇)，12000rpm離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)1min，棄廢液(ye)(ye)，將吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)入(ru)(ru)(ru)(ru)(ru)收(shou)集(ji)管(guan)中(zhong)(zhong)(zhong)。6、向(xiang)吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)500ul漂(piao)洗(xi)(xi)液(ye)(ye)，12000rpm離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)1min，棄廢液(ye)(ye)，將吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)入(ru)(ru)(ru)(ru)(ru)收(shou)集(ji)管(guan)中(zhong)(zhong)(zhong)。7、 12000rpm離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)2min，將吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)置(zhi)于室(shi)溫(wen)或50℃溫(wen)箱放(fang)置(zhi)數(shu)分(fen)鐘，目(mu)的(de)(de)是將吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)殘余(yu)的(de)(de)漂(piao)洗(xi)(xi)液(ye)(ye)去除， 否則漂(piao)洗(xi)(xi)液(ye)(ye)中(zhong)(zhong)(zhong)的(de)(de)乙(yi)醇會(hui)(hui)影響(xiang)后續的(de)(de)實驗如酶切(qie)、PCR等。8、將吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)入(ru)(ru)(ru)(ru)(ru)一(yi)個(ge)干凈的(de)(de)離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)管(guan)中(zhong)(zhong)(zhong)，向(xiang)吸(xi)(xi)附(fu)(fu)(fu)膜(mo)中(zhong)(zhong)(zhong)央懸空滴(di)加(jia)(jia)(jia)(jia)(jia)50ul-100ul經65℃水浴預熱的(de)(de)洗(xi)(xi)脫液(ye)(ye)，室(shi)溫(wen)放(fang)置(zhi)5min，12000rpm離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)1min。9、離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)所得(de)(de)洗(xi)(xi)脫液(ye)(ye)再加(jia)(jia)(jia)(jia)(jia)入(ru)(ru)(ru)(ru)(ru)吸(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)，室(shi)溫(wen)放(fang)置(zhi)2min，12000rpm離(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)2min，即(ji)可(ke)得(de)(de)到高質量(liang)的(de)(de)病毒(du)基(ji)因(yin)組DNA。

solarbio-DNA病毒基因組提取試劑盒

注意事項:
1、試劑盒拆封后，蛋白酶K需放置-20℃保存。
2、樣品應避免反復凍融，否則會導致提取的DNA段較小且提取量也下降。
3、若結合液中有沉淀，可在37℃水浴中重新溶解再使用，不影響效果。
4、洗脫緩沖液的體積好不少于50ul，體積過小會影響回收效率。洗脫液的pH值對洗脫效率也有影響，若需要用水做洗脫液應保證其pH值在8.0左右(可用NaOH將水的pH值調此范圍)，pH 值低于7.0會降低洗脫效率。DNA產物應保存在-20℃，以防DNA降解。
5、DNA濃度(du)及純(chun)度(du)檢測：得到(dao)的(de)基因組(zu)DNA段的(de)大小與(yu)病毒(du)的(de)保存條件和(he)種(zhong)類等因素有(you)關。D260值(zhi)(zhi)為(wei)(wei)1.0相(xiang)當于大約50 ug/ml雙鏈DNA、40 ug/ml單鏈DNA。OD260/OD280比值(zhi)(zhi)應為(wei)(wei)1.7-1.9，如果洗脫時不使用洗脫緩沖液，而使用去離子水，比值(zhi)(zhi)會偏(pian)低，因為(wei)(wei)pH值(zhi)(zhi)和(he)離子存在會影響光吸收值(zhi)(zhi)，但并不表(biao)示純(chun)度(du)低。6、如果病毒(du)含(han)量過低，后提取的(de)基因組(zu)DAN電泳可(ke)能無(wu)法檢測到(dao)，但PCR等其他(ta)實驗還會有(you)結果。

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