本文選題：細(xì)胞交叉污染 + 細(xì)胞身份鑒定��； 參考：《武漢大學(xué)》2017年博士論文

【摘要】：細(xì)胞系作為生命科學(xué)、臨床醫(yī)學(xué)等研究的常用材料,其重要性不言而喻,但細(xì)胞的質(zhì)量鑒定問題一直被許多研究者所忽視,細(xì)胞污染和細(xì)胞身份誤判的現(xiàn)象非常普遍。近年來很多權(quán)威期刊和研究機(jī)構(gòu)建議應(yīng)該在研究論文出版發(fā)行或是申請(qǐng)基金之前進(jìn)行相關(guān)細(xì)胞系的鑒定。雖然現(xiàn)在有很多新的方法能對(duì)細(xì)胞進(jìn)行鑒定,但是基于短串聯(lián)重復(fù)序列的DNA分型技術(shù)仍然是作為細(xì)胞鑒定的金標(biāo)準(zhǔn)。短串聯(lián)重復(fù)序列(Short Tandom Repeat,STR)是由2-7個(gè)堿基對(duì)作為核心單位串聯(lián)重復(fù)形成的一類DNA序列。由于具有廣泛分布,信息量大,高度多態(tài)性并遵循孟德爾遺傳規(guī)律,易于PCR擴(kuò)增及分型等特征,STR技術(shù)已廣泛用于親子鑒定及人源細(xì)胞系交叉污染檢測(cè)和身份鑒定。本論文利用21位點(diǎn)的STR分型新技術(shù)進(jìn)行人源細(xì)胞系交叉污染檢測(cè)和身份鑒定,共檢測(cè)了來自28個(gè)研究單位的278株細(xì)胞系。檢測(cè)結(jié)果發(fā)現(xiàn),在278株細(xì)胞系中,46%的細(xì)胞存在交叉污染,Hep2和EJ細(xì)胞系還在錯(cuò)誤身份下使用。73%(71株中52株)的交叉污染細(xì)胞來自于中國,占了交叉污染細(xì)胞總數(shù)的40%(129株中52株)。67%的污染源是HeLa細(xì)胞或者是HeLa細(xì)胞與其他細(xì)胞相互雜交的亞細(xì)胞株。肝癌細(xì)胞系HCCC-9810和退行性的肺癌細(xì)胞系Calu-6用9個(gè)位點(diǎn)的STR圖譜數(shù)據(jù)(ATCC STR數(shù)據(jù)庫)比對(duì)分析,顯示了 0.89的相似性,而0.89的比對(duì)數(shù)據(jù)提示這兩株細(xì)胞是同源細(xì)胞系。21位點(diǎn)的STR檢測(cè)圖譜分析和24個(gè)位點(diǎn)的SNP(Single Nucleotide Polymorphisms,SNP)檢測(cè)結(jié)果均證明兩株細(xì)胞是非同源細(xì)胞系,證明用21位點(diǎn)的STR新技術(shù)鑒定人源細(xì)胞系,數(shù)據(jù)更精確,結(jié)果更可靠。150株正確的細(xì)胞系展示了獨(dú)特的分型圖譜,排除了交叉污染,確認(rèn)了身份。通過構(gòu)建自主知識(shí)產(chǎn)權(quán)的細(xì)胞系STR數(shù)據(jù)庫,為國內(nèi)人源細(xì)胞系的STR分型提供比對(duì)標(biāo)準(zhǔn),具有重要現(xiàn)實(shí)意義。
[Abstract]:The importance of cell line as a common material in life science and clinical medicine is self-evident, but the problem of cell quality identification has been ignored by many researchers. The phenomenon of cell contamination and misjudgment of cell identity is very common. In recent years, many leading journals and research institutions have suggested that cell lines should be identified before research papers are published or funds are applied. Although there are many new methods for cell identification, DNA typing based on short tandem repeats is still the golden standard for cell identification. Short Tandom repeat sequence (STR) is a kind of DNA sequence which is composed of 2-7 base pairs as core unit tandem repeats. Due to its wide distribution, large amount of information, high polymorphism and Mendelian genetic rule, PCR amplification and typing have been widely used in paternity testing, human cell line cross-contamination detection and identification. In this paper, a new 21 locus STR typing technique was used to detect the cross contamination and identification of human cell lines. A total of 278 cell lines from 28 research units were detected. The results showed that 46% of the 278 cell lines contained cross contamination of Hep2 and EJ cell lines. 52 or 67% of the total number of cross-polluted cells were HeLa cells or sub-cell lines of HeLa cells hybridizing with other cells. HCCC-9810 and Calu-6, a degenerative lung cancer cell line, were compared with each other by STR map data of 9 loci (ATCC STR database). The comparison data of 0.89 and 0.89 showed that the two cell lines were non-homologous cell lines. The results of STR analysis and 24 loci SNP(Single Nucleotide PolymorphismsSNPs analysis showed that the two cell lines were non-homologous. It is proved that the new technique of STR at 21 loci is more accurate in identifying human cell lines, and the results are more reliable. 150 strains of correct cell lines exhibit unique genotyping patterns, eliminate cross contamination and confirm their identities. It is of great practical significance to construct the STR database of human cell line, which can provide a comparison standard for STR typing of human cell lines in China.
【學(xué)位授予單位】：武漢大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：R329.2

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本文編號(hào)：1924914