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細胞STR鑒定簡介:

細胞STR鑒定應用于生物醫學研究領域的哺乳動物細胞被錯誤鑒定和交叉污染的問題,一直是一個普遍存在的突出問題。

據統計,國外實驗室有20%左右的細胞株被錯誤鑒定和交叉污染,NIH和ATCC近兩年都對此發出呼吁,要求研究者對細胞進行鑒定。近年來,大量研究表明STR基因分型方法是進行細胞交叉污染和性質鑒定的最有效和準確的方法之一,STR基因分型應用于細胞鑒定已被ATCC等機構強烈推薦。美國的ATCC細胞庫、德國的DSMZ細胞庫以及日本的JCRB細胞庫等為STR分型提供了各細胞株的數據供比對。

STR(Short Tandem Repeat,短串聯重復序列)基因位點由長度為3-7個堿基對的短串連重復序列組成,這些重復序列廣泛存在于人類基因組中,可作為高度多態性標記,被稱為細胞的DNA指紋,其可通過PCR(聚合酶鏈式反應)來檢測。STR基因座位上的等位基因可通過擴增區域內重復序列的拷貝數的不同來區分,在毛細管電泳分離之后可通過熒光檢測來識別。隨后通過一定的計算方法,即可根據所得的STR分型結果與專業的細胞STR數據庫比對從而推算出樣品所屬的細胞系或可能的交叉污染的細胞系名稱。


細胞STR鑒定服務流程:

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細胞STR鑒定樣本要求:

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細胞系錯誤鑒定所造成的后果:

★ 細胞系的損失;

★ 時間和金錢的損失;

★ 在公眾領域(文獻、專利等)提供錯誤信息;

★ 造成實驗結果不一致或不可重復;

★ ……


何時需做細胞STR鑒定?

1、發表文章或申請課題經費前;

2、使用細胞進行臨床治療(試驗)前;

3、準備凍存保種或已凍存多年的細胞;

4、一個涉及到細胞試驗項目開始/結束時;

5、新得到的細胞或實驗室傳5代以上的細胞;

6、細胞系表現不穩定或結果與預期差別較大。


已開始要求細胞鑒定的期刊:

★ Nature

★ BioTechniques

★ Cancer Research

★ Cancer Discovery

★ Clinical Cancer Research

★ Molecular Cancer Research

★ Cancer Prevention Research

★ International Journal of Cancer

★ Molecular Cancer Therapeutics

★ Cell Biochemistry and Biophysics

★ Cancer Epidemiology, Biomarkers & Prevention

★ In Vitro Cellular & Developmental Biology - Animal

★ ......


人源細胞STR位點信息:

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為進一步提高鑒定的分辨率,除了包含ATCC檢測的8個STR和1個性別決定位點Amelogenin外,另新增了12個高度多態性位點,共檢測21個STR位點。


參考文獻:

1. Chatterjee, R. (2007) Cell biology. Cases of mistaken identity. Science 315, 928-31.

2. Ruiz Bravo, N. and Gottesman, M. (2007) Notice regarding authentication of cultured cell lines.

3. Yoshino, K. et al. (2006) Essential role for gene profiling analysis in the authentication of human cell lines. Human Cell 19, 43-8.

4. Szibor, R. et al. (2003) Cell line DNA typing in forensic genetics—the necessity of reliable standards. Forensic Sci. Int. 138, 37-43.

5. Dirks, W.G. et al. (2005) Short tandem repeat DNA typing provides an international reference standard for authentication of human cell lines. ALTEX 22, 103-9.

6. Masters, J.R. et al. (2001) Short tandem repeat profiling provides an international reference standard for human cell lines. Proc. Natl. Acad. Sci. USA 98, 8012-7.

7. (2001) Verify cell line identity with DNA profiling. ATCC Connection: Newsletter of The American Type Culture Collection 21, 1-2.

8. Krenke, B. et al. (2002) Validation of a 16-locus fluorescent multiplex system. J. Forensic Sci. 47, 773-85.

9. Levinson, G. and Gutman, G.A. (1987) Slipped-strand mispairing: A major mechanism for DNA sequence evolution. Mol. Biol. Evol. 4, 203-21.

10. Schlotterer, C. and Tautz, D. (1992) Slippage synthesis of simple sequence DNA. Nucleic Acids Res. 20, 211-5.

11. Smith, J.R. et al. (1995) Approach to genotyping errors caused by nontemplated nucleotide addition by Taq DNA polymerase. Genome Res. 5, 312-7.

12. Magnuson, V.L. et al. (1996) Substrate nucleotide-determined non-templated addition of adenine by Taq DNA polymerase: Implications for PCR-based genotyping. BioTechniques 21, 700-9.

13. Walsh, P.S., Fildes, N.J. and Reynolds, R. (1996) Sequence analysis and characterization of stutter products at the tetranucleotide repeat locus vWA. Nucleic Acids Res. 24, 2807-12.

14. Moller, A., Meyer, E. and Brinkmann, B. (1994) Different types of structural variation in STRs: HumFES/FPS, HumVWA and HumD21S11. Int. J. Leg. Med. 106, 319-23.

15. Brinkmann, B., Moller A. and Wiegand, P. (1995) Structure of new mutations in 2 STR systems. Int. J. Leg. Med. 107, 201-3.

16. Griffiths, R. et al. (1998) New reference allelic ladders to improve allelic designation in a multiplex STR system. Int. J. Legal Med. 111, 267-72.

17. B?r, W. et al. (1997) DNA recommendations: Further report of the DNA Commission of the ISFH regarding the use of short tandem repeat systems. Int. J. Legal Med. 110, 175-6.

18. Gill, P. et al. (1997) Considerations from the European DNA Profiling Group (EDNAP) concerning STR nomenclature. Forensic Sci. Int. 87, 185-92.

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