【摘要】：為建立犬細(xì)環(huán)病毒(TTCV)的快速檢測方法,本研究根據(jù)GenBank中登錄的TTCV ORF7基因設(shè)計(jì)合成一對特異性引物,并對反應(yīng)條件和反應(yīng)體系進(jìn)行優(yōu)化,建立了檢測TTCV的SYBR Green I熒光定量PCR方法。結(jié)果顯示:建立的熒光定量PCR方法 Ct值與標(biāo)準(zhǔn)品模板在5.82×10~9拷貝/μL~5.82×10~3拷貝/μL范圍內(nèi)呈良好的線性關(guān)系。該檢測方法僅對TTCV檢測為陽性,而對狂犬病病毒、犬冠狀病毒、犬細(xì)小病毒、犬瘟熱病毒和犬副流感病毒均無特異擴(kuò)增;該檢測方法靈敏度可達(dá)5.82×103拷貝/μL。對27份犬血清樣品檢測結(jié)果顯示,建立的熒光定量PCR陽性檢測率為11.11%,而普通PCR方法檢測陽性率為3.7%。本實(shí)驗(yàn)建立了TTCV SYBR Green Ⅰ熒光定量PCR檢測方法,對TTCV診斷及致病機(jī)制的研究提供了高通量的定量檢測方法。
[Abstract]:In order to establish a rapid detection method for (TTCV) of canine fine loop virus, a pair of specific primers were designed and synthesized according to the TTCV ORF7 gene registered in GenBank, and the reaction conditions and reaction system were optimized. A SYBR Green I fluorescence quantitative PCR method for the detection of TTCV was established. The results showed that there was a good linear relationship between the Ct value and the standard sample template in the range of 5.82 脳 10 ~ (9) copies / 渭 L ~ 5.82 脳 10 ~ (-3) copies / 渭 L. The sensitivity of this method was 5.82 脳 10 ~ 3 copies / 渭 L for rabies virus, canine coronavirus, canine parvovirus, canine distemper virus and canine parainfluenza virus. The positive rate of fluorescent quantitative PCR was 11.11% in 27 canine serum samples, while the positive rate of normal PCR method was 3.7%. In this study, a fluorescence quantitative PCR assay for the detection of TTCV SYBR Green 鈪，

本文編號：2376426