本文選題：緩沖液　切入點(diǎn)：全血擴(kuò)增　出處：《中國(guó)新藥雜志》2016年24期

【摘要】：目的:研制一種以全血和濾紙干血為模板的等位基因特異性PCR(allele-specific polymerase chain reaction,AS-PCR)體系的緩沖液。方法:采用血樣直接作為模板,通過引入錯(cuò)配堿基對(duì)AS-PCR的特異性進(jìn)行改進(jìn),研制一種新型緩沖液HPEC[p H 9.5,Mg2+3.5 mmol·L~(-1),海藻糖2.5μmol·L~(-1),(NH4)2SO49.6mmol·L~(-1),Tween-20 0.01%],對(duì)包含單核苷酸多態(tài)性位點(diǎn)(single nucleotide polymorphisms,SNPs)的目的片段進(jìn)行擴(kuò)增。結(jié)果:該方法成功地實(shí)現(xiàn)了以血樣和濾紙干血為模板的AS-PCR基因分型。結(jié)論:此緩沖液省去了擴(kuò)增前基因組的提取和純化,簡(jiǎn)化了操作過程,降低DNA污染機(jī)會(huì)和檢測(cè)技術(shù)的復(fù)雜性。
[Abstract]:Objective: to develop a buffer solution of allele-specific PCR(allele-specific polymerase chain reactionsis-AS-PCR system with whole blood and filter paper dry blood as template. Methods: the specificity of AS-PCR was improved by introducing mismatch base. A novel buffer solution, HPEC [p H 9.5 mg 23. 5 mmol / L HPEC, trehalose 2. 5 渭 mol L ~ (-1)), NH _ 4N _ 2SO _ 4 9. 6 mmol / L ~ (-1) ~ (-1)] was developed to amplify the target fragment containing single nucleotide polymorphisms (SNPs). Results: this method successfully realized the AS-PCR base using blood samples and filter paper dry blood as templates. Conclusion: this buffer eliminates the extraction and purification of the genome before amplification. It simplifies the operation process, reduces the chance of DNA pollution and the complexity of detection technology.
【作者單位】： 解放軍211醫(yī)院;華僑大學(xué)生物醫(yī)學(xué)學(xué)院;
【基金】：福建省自然科學(xué)基金資助項(xiàng)目(2015J01342)
【分類號(hào)】：R440
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本文編號(hào)：1667795