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solarbio-DNA病毒基因組提取試劑盒
本(ben)試(shi)劑盒適合于從血清、細胞上(shang)清、淋(lin)巴液(ye)中(zhong)(zhong)(zhong)提(ti)(ti)取(qu)DNA病毒(du)(du)基(ji)因(yin)組(zu)，不(bu)適合于RNA病毒(du)(du)基(ji)因(yin)組(zu)的(de)提(ti)(ti)取(qu)。使(shi)用(yong)(yong)本(ben)試(shi)劑盒提(ti)(ti)取(qu)的(de)基(ji)因(yin)組(zu)DNA可(ke)(ke)用(yong)(yong)于各種常(chang)規操作，包括酶(mei)(mei)切、PCR、文庫構建、Southern雜交等實驗。操作步(bu)(bu)驟： 使(shi)用(yong)(yong)前請(qing)先在(zai)漂(piao)洗(xi)(xi)液(ye)中(zhong)(zhong)(zhong)加(jia)入(ru)(ru)(ru)無水(shui)乙(yi)醇，加(jia)入(ru)(ru)(ru)體(ti)積請(qing)參照(zhao)瓶上(shang)的(de)標簽。所有離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)步(bu)(bu)驟均為使(shi)用(yong)(yong)臺式離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)機(ji)在(zai)室溫下離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)。1、取(qu)病毒(du)(du)上(shang)清液(ye)0.5ml，12000rpm離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)5min，盡量吸(xi)(xi)(xi)盡上(shang)清使(shi)用(yong)(yong)，棄去沉(chen)淀。2、向(xiang)病毒(du)(du)上(shang)清中(zhong)(zhong)(zhong)加(jia)入(ru)(ru)(ru)20ul 10mg/ml的(de)蛋白酶(mei)(mei)K，充分混勻，65℃消化10-20min，期間可(ke)(ke)顛倒離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)管混勻數次(ci)(ci)。3、向(xiang)管中(zhong)(zhong)(zhong)加(jia)入(ru)(ru)(ru)500ul結合液(ye)，充分混勻。再向(xiang)管中(zhong)(zhong)(zhong)加(jia)入(ru)(ru)(ru)400ul無水(shui)乙(yi)醇，充分混勻，此時可(ke)(ke)能會出現絮狀沉(chen)淀，不(bu)影響DNA的(de)提(ti)(ti)取(qu)，可(ke)(ke)將(jiang)(jiang)(jiang)溶液(ye)和絮狀沉(chen)淀都加(jia)入(ru)(ru)(ru)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)，靜置(zhi)2min。（吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)的(de)大(da)容積為750ul，可(ke)(ke)分兩次(ci)(ci)加(jia)入(ru)(ru)(ru)。一次(ci)(ci)吸(xi)(xi)(xi)附(fu)(fu)(fu)完(wan)離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)后(hou)再將(jiang)(jiang)(jiang)余下的(de)混合液(ye)體(ti)加(jia)入(ru)(ru)(ru)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)靜置(zhi)離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)。） 4、 12000rpm離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)2min，棄廢液(ye)，將(jiang)(jiang)(jiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)(fang)(fang)(fang)入(ru)(ru)(ru)收集管中(zhong)(zhong)(zhong)。5、向(xiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)加(jia)入(ru)(ru)(ru)700ul漂(piao)洗(xi)(xi)液(ye)(使(shi)用(yong)(yong)前請(qing)先檢查是(shi)否(fou)已(yi)加(jia)入(ru)(ru)(ru)無水(shui)乙(yi)醇)，12000rpm離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)1min，棄廢液(ye)，將(jiang)(jiang)(jiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)(fang)(fang)(fang)入(ru)(ru)(ru)收集管中(zhong)(zhong)(zhong)。6、向(xiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)加(jia)入(ru)(ru)(ru)500ul漂(piao)洗(xi)(xi)液(ye)，12000rpm離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)1min，棄廢液(ye)，將(jiang)(jiang)(jiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)(fang)(fang)(fang)入(ru)(ru)(ru)收集管中(zhong)(zhong)(zhong)。7、 12000rpm離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)2min，將(jiang)(jiang)(jiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)置(zhi)于室溫或50℃溫箱放(fang)(fang)(fang)(fang)置(zhi)數分鐘，目的(de)是(shi)將(jiang)(jiang)(jiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)殘余的(de)漂(piao)洗(xi)(xi)液(ye)去除， 否(fou)則漂(piao)洗(xi)(xi)液(ye)中(zhong)(zhong)(zhong)的(de)乙(yi)醇會影響后(hou)續的(de)實驗如酶(mei)(mei)切、PCR等。8、將(jiang)(jiang)(jiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)放(fang)(fang)(fang)(fang)入(ru)(ru)(ru)一個干凈的(de)離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)管中(zhong)(zhong)(zhong)，向(xiang)吸(xi)(xi)(xi)附(fu)(fu)(fu)膜中(zhong)(zhong)(zhong)央懸空滴加(jia)50ul-100ul經65℃水(shui)浴預熱的(de)洗(xi)(xi)脫液(ye)，室溫放(fang)(fang)(fang)(fang)置(zhi)5min，12000rpm離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)1min。9、離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)所得(de)洗(xi)(xi)脫液(ye)再加(jia)入(ru)(ru)(ru)吸(xi)(xi)(xi)附(fu)(fu)(fu)柱(zhu)(zhu)(zhu)中(zhong)(zhong)(zhong)，室溫放(fang)(fang)(fang)(fang)置(zhi)2min，12000rpm離(li)(li)(li)(li)(li)(li)(li)心(xin)(xin)(xin)(xin)2min，即可(ke)(ke)得(de)到高質量的(de)病毒(du)(du)基(ji)因(yin)組(zu)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濃度及純(chun)度檢測(ce)：得到(dao)的基(ji)因組DNA段的大小(xiao)與病毒的保(bao)存(cun)條件和種類等因素有關。D260值為1.0相當(dang)于大約50 ug/ml雙(shuang)鏈DNA、40 ug/ml單鏈DNA。OD260/OD280比(bi)值應為1.7-1.9，如果洗(xi)脫時(shi)不(bu)使(shi)用洗(xi)脫緩沖液，而使(shi)用去離(li)子水，比(bi)值會偏低，因為pH值和離(li)子存(cun)在會影(ying)響光(guang)吸收值，但并不(bu)表示純(chun)度低。6、如果病毒含量過(guo)低，后提取(qu)的基(ji)因組DAN電泳可能無(wu)法檢測(ce)到(dao)，但PCR等其他(ta)實驗還(huan)會有結果。

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