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瓊脂糖水平電泳儀:什么是核酸電泳?

發(fa)布(bu)時(shi)間:2022-06-22

  瓊(qiong)脂糖(tang)水平(ping)電(dian)(dian)泳(yong)(yong)儀:什么是(shi)(shi)核(he)酸(suan)(suan)電(dian)(dian)泳(yong)(yong)?  核(he)酸(suan)(suan)電(dian)(dian)泳(yong)(yong)通(tong)常(chang)使(shi)(shi)用(yong)(yong)(yong)(yong)瓊(qiong)脂糖(tang)凝(ning)膠(jiao)(jiao)和聚(ju)(ju)(ju)丙(bing)(bing)烯(xi)(xi)(xi)酰(xian)(xian)胺(an)(an)(an)凝(ning)膠(jiao)(jiao)兩種(zhong)(zhong)介質。瓊(qiong)脂糖(tang)是(shi)(shi)一(yi)種(zhong)(zhong)聚(ju)(ju)(ju)合(he)鏈線性分(fen)子;聚(ju)(ju)(ju)丙(bing)(bing)烯(xi)(xi)(xi)酰(xian)(xian)胺(an)(an)(an)凝(ning)膠(jiao)(jiao)由丙(bing)(bing)烯(xi)(xi)(xi)酰(xian)(xian)胺(an)(an)(an)(Acr)在(zai)(zai) N,N,N′-四(si)甲(jia)基(ji)乙(yi)(yi) 二胺(an)(an)(an)(TEMED)和過(guo)硫酸(suan)(suan)銨(AP)的(de)催化下聚(ju)(ju)(ju)合(he)形成(cheng)長鏈,并(bing)通(tong)過(guo)交聯劑 N,N′-亞甲(jia)雙丙(bing)(bing)烯(xi)(xi)(xi)酰(xian)(xian)胺(an)(an)(an)(Bis)交叉(cha)連接而(er)成(cheng)。瓊(qiong)脂糖(tang)凝(ning)膠(jiao)(jiao)適合(he)分(fen)離(li)200bp至近50kb的(de)DN 片(pian)(pian)段(duan),而(er)聚(ju)(ju)(ju)丙(bing)(bing)烯(xi)(xi)(xi)酰(xian)(xian)胺(an)(an)(an)凝(ning)膠(jiao)(jiao)對于小片(pian)(pian)段(duan)(5bp-500bp)的(de)分(fen)離(li)效果(guo)很好(hao),且(qie)分(fen)辯力極高。選(xuan)擇不同(tong)濃度的(de)凝(ning)膠(jiao)(jiao),可(ke)以(yi)分(fen)離(li)丌同(tong)大(da)(da)小范(fan)圍(wei)的(de) DNA 分(fen)子。電(dian)(dian)場強度愈大(da)(da),帶電(dian)(dian)顆粒的(de)運動赹(qiong)快。但(dan)凝(ning)膠(jiao)(jiao)的(de)有效分(fen)離(li)范(fan)圍(wei)隨(sui)著電(dian)(dian)壓增大(da)(da)而(er)減小,所(suo)以(yi)電(dian)(dian)泳(yong)(yong)時一(yi)般采(cai)用(yong)(yong)(yong)(yong)低(di)電(dian)(dian)壓,不超過(guo) 6V/cm。而(er)對于大(da)(da)片(pian)(pian)段(duan)電(dian)(dian)泳(yong)(yong),可(ke)以(yi)選(xuan)擇 0.5-1.0V/cm 電(dian)(dian)泳(yong)(yong)過(guo)夜(ye)。如果(guo)選(xuan)擇高電(dian)(dian)壓電(dian)(dian)泳(yong)(yong),可(ke)使(shi)(shi)用(yong)(yong)(yong)(yong)聚(ju)(ju)(ju)丙(bing)(bing)烯(xi)(xi)(xi)酰(xian)(xian)胺(an)(an)(an)凝(ning)膠(jiao)(jiao)。  核(he)酸(suan)(suan)電(dian)(dian)泳(yong)(yong)常(chang)采(cai)用(yong)(yong)(yong)(yong)TAE、TBE、TPE三種(zhong)(zhong)緩(huan)沖系統。TAE價格低(di)廉,但(dan)緩(huan)沖能力低(di)。TPE在(zai)(zai)進行DNA回收時,會使(shi)(shi)DNA污染磷酸(suan)(suan)鹽,影響(xiang)后續反(fan)應。所(suo)以(yi)綜合(he)考慮,多采(cai)用(yong)(yong)(yong)(yong)TBE緩(huan)沖液(ye)。  核(he)酸(suan)(suan)電(dian)(dian)泳(yong)(yong)中常(chang)用(yong)(yong)(yong)(yong)的(de)染色劑是(shi)(shi)溴化乙(yi)(yi)錠(EB)。溴化乙(yi)(yi)錠是(shi)(shi)一(yi)種(zhong)(zhong)扁平(ping)分(fen)子,可(ke)以(yi)嵌入核(he)酸(suan)(suan)雙鏈的(de)配(pei)對堿基(ji)之間。

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