嗜热链球菌中CRISPR序列的检测与同源性分析

食品科学 ›› 2013, Vol. 34 ›› Issue (3): 153-157.

嗜热链球菌中CRISPR序列的检测与同源性分析

邓凯波¹,霍贵成²

1. 东北农业大学食品学院乳品科学教育部重点实验室
2. 乳品科学教育部重点实验室（东北农业大学）

收稿日期:2011-10-28 修回日期:2013-01-05 出版日期:2013-02-15 发布日期:2017-12-29
通讯作者: 邓凯波 E-mail:keppel_red@yahoo.com.cn
基金资助:
国家自然科学基金面上项目;教育部长江学者和创新团队发展计划

Detection and Homologous Analysis of CRISPR in Streptococcus thermophilus

Received:2011-10-28 Revised:2013-01-05 Online:2013-02-15 Published:2017-12-29
Contact: Kai-Bo DENG E-mail:keppel_red@yahoo.com.cn

摘要/Abstract

摘要： CRISPR为近年发现的一种广泛存在于原核生物中的获得性免疫系统。为探明内蒙古的8株嗜热链球菌的CRISPR，用PCR方法扩增了其CRISPR序列，并采用生物信息学方法分析和预测了重复序列（Direct Repeat，DR）的同源性及二级结构。结果表明，除S4只含有一个CRISPR结构外，其余7株嗜热链球菌均检测出3条CRISPR序列，远高于CRISPR database公布细菌中具有该结构的比例（46.4%），且分别具特异性的重复序列（DR1-DR3）；CRISPR最长达2853 bp，最短仅101 bp。对DR的二级结构预测发现，DR1-DR3均能形成回文结构，但茎环大小会有差异；在同源性比对中发现，除多数同属或同种外，供试DR还与个别远缘菌种具有高同源性，这种现象证明了供试菌株的CRISPR序列同样存在的水平基因转移，并可能存在其他的进化进程。

Abstract: CRISPR was recently found to be an acquired immune system widespread in prokaryotes. In order to find out CRISPRs in 8 strains of Streptococcus thermophilus isolated from Inner Mongolia, the CRISPR sequences were located by PCR amplification, and homology and secondary structure of direct repeat (DR) were predicted using bioinformatics method. Results showed that all the studied strains contained three CRISPR sequences except one in S4, which was much higher than the ratio (46.4%) in bacteria published on CRISPR database, and three CRISPRs had their specialized direct repeats (DR1-DR3)；The longest CRISPR was 2853 bp, and the shortest was only 101 bp. According to the prediction of secondary structure, a palindrome were detected in DR1-DR3, respectively, although the sizes of stem-loop were different; for the homology analysis, DR demonstrated high similarity with several individual species of distant relationships like those in the same genus or species, this suggested that the CRISPRs of tested strains also possess the characteristic of horizontal gene transfer (HGT), as well as the possibility of other evolutionary process.

中图分类号:

邓凯波霍贵成. 嗜热链球菌中CRISPR序列的检测与同源性分析[J]. 食品科学, 2013, 34(3): 153-157.

参考文献

[1] STURINO JM and KLAENHAMMER TR. Engineered Bacteriophage-defence Systems in Bioprocessing[J]. Nature, 2006, 4(5): 395-404;
[2] JANSEN R, EMBDEN JD, GAASTRA W, et al. Identification of Genes that are Associated with DNA Repeats in Prokaryotes[J]. Mol Microbiol, 2002, 43 (6): 1565-1575;
[3] SOREK R, KUNIN V, HUGENHOLTZ P. CRISPR-a Widespread System that Provides Acquired Resistance against Ohages in Bacteria and Archaea[J]. Nat Rev Microbiol, 2008, 6(3): 181-186;
[4] BARRANGOU R, FREMAUX C, DEVEAU H, et al. CRISPR Provides Acquired Resistance against Viruses in Prokaryotes[J]. Science, 2007, 315 (5819): 1709-1712;
[5] BARRANGOU R and HORVATH P. The CRISPR System Protects Microbes against Phages, Plasmids[J]. Microbe, 2009, 4(5): 224-230;
[6] DEVEAU H, BARRANGOU R, GARNEAU J E, et al. 2008. Phage Response to CRSIPR-encoded Resistance in Streptococcus thermophilus[J]. J Bacteriol, 190: 1390-1400;
[7] HORVATH P, ROMERO D A, CO?Té-MONVOISIN A, et al. Diversity, Activity and Evolution of CRISPR Loci in Streptococcus thermophilus[J]. J Bacteriol, 2008, 190(4): 1401-1412;
[8] GARNEAU JE, Dupuis Mè, Villion M, et al. The CRISPR/Cas Bacterial Immune System Cleaves Bacteriophage and Plasmid DNA[J]. Nature, 2010, 468(7320): 67-71;
[9] BOLOTIN A, QUINQUIS B, SOROKIN A, et al. Clustered Regularly Interspaced Short Palindrome Repeats (CRISPRs) Have Spacers of Extrachromosomal Origin [J]. Microbiology, 2005, 151(8): 2551-2561;
[10] GODDE JS and BICHERTON A. The Repetitive DNA Elements Called CRISPRs and Their Associated Genes: Evidence of Horizontal Rransfer among Prokaryotes[J]. J Mol Evol, 2006, 62(6): 718-729;
[11] MARRAFFINI LA, and SONTHEIMER E J. CRISPR Interference Limits Horizontal Gene Transfer in Staphylococci by Targeting DNA[J]. Science, 2008, 322(5909): 1843-1845;
[12] LIU Mengjin, SIEZEN R J and NAUTA A. In Silico Prediction of Horizontal Gene Transfer Events in Lactobacillus bulgaricus and Streptococcus thermophilus Reveals Protocooperation in Yogurt Manufacturing[J]. AEM, 2009, 75(12): 4120-4129;
[13] HORVATH P, CO?Té-MONVOISIN A, ROMERO DA, et al. Comparative Analysis of CRISPR Loci in Lactic Acid Bacteria Genomes[J]. Int J Food Microbiol, 2009, 131(1): 62-70;
[14] CHAKRABORTY S, SNIJDERS AP, CHAKRAVORTY R, et al. Comparative Network Clustering of Direct Repeats (DRs) and cas genes Confirms the Possibility of the Horizontal Transfer of CRISPR Locus among Bacteria[J]. Mol Phylogenet Evol, 2010, 56(3): 878-887;
[15] POUECEL C, SALVIGNOL G and VERGNAUD G. CRISPR Elements in Yersinia pestis Acquire New Repeats by Preferential Uptake of Bacteriophage DNA, and Provide Additional Tools for Evolutionary Studies[J]. Microbiology, 2005, 151(3): 653-663;