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靶向鉴别、检测动物双歧杆菌的引物探针及应用

杜丽霞, 李典, 禚惠荣, 石梦楠, 陈延宁, 李岚芳, 侯少阳

杜丽霞, 李典, 禚惠荣, 石梦楠, 陈延宁, 李岚芳, 侯少阳. 靶向鉴别、检测动物双歧杆菌的引物探针及应用[J]. 中国药科大学学报, 2023, 54(2): 255-262. DOI: 10.11665/j.issn.1000-5048.20220912001
引用本文: 杜丽霞, 李典, 禚惠荣, 石梦楠, 陈延宁, 李岚芳, 侯少阳. 靶向鉴别、检测动物双歧杆菌的引物探针及应用[J]. 中国药科大学学报, 2023, 54(2): 255-262. DOI: 10.11665/j.issn.1000-5048.20220912001
DU Lixia, LI Dian, ZHUO Huirong, SHI Mengnan, CHEN Yanning, LI Lanfang, HOU Shaoyang. Primer probes for targeted identification and detection of Bifidobacterium animalis and their application[J]. Journal of China Pharmaceutical University, 2023, 54(2): 255-262. DOI: 10.11665/j.issn.1000-5048.20220912001
Citation: DU Lixia, LI Dian, ZHUO Huirong, SHI Mengnan, CHEN Yanning, LI Lanfang, HOU Shaoyang. Primer probes for targeted identification and detection of Bifidobacterium animalis and their application[J]. Journal of China Pharmaceutical University, 2023, 54(2): 255-262. DOI: 10.11665/j.issn.1000-5048.20220912001

靶向鉴别、检测动物双歧杆菌的引物探针及应用

基金项目: 山东省自然科学基金资助项目(No.ZR2022QB146);菏泽学院博士基金资助项目(No.XY21BS35)

Primer probes for targeted identification and detection of Bifidobacterium animalis and their application

Funds: This study was supported by the Natural Science Foundation of Shandong Province (No.ZR2022QB146); and the Heze University PhD Fund Planning (No.XY21BS35)
  • 摘要: 为解决目前应用于鉴别动物双歧杆菌的方法的用时长、操作复杂、实验环境低适配性等缺点,开发了一种基于ERIC-PCR技术设计特异性引物探针以靶向鉴别、检测动物双歧杆菌的方法。以动物双歧杆菌HP-B1124基因组DNA为基础,优化动物双歧杆菌HP-B1124的ERIC-PCR反应条件,对ERIC-PCR片段逐一回收并进行测序,针对测序结果设计两对特异性引物探针,对两对引物探针设计结果的准确性、特异性、可检测基因组DNA的含量限度、普适性进行检验,并运用所设计的两对特异性引物探针检验市售公示配方中含有动物双歧杆菌的产品。本研究设计的两对特异性引物探针可简单、快速、靶向的用于动物双歧杆菌的鉴定。此方法优化了目前相对传统的动物双歧杆菌纯培养及平板计数鉴定方法,一定程度上解决了SNP基因分型技术及实时荧光定量PCR法等方法在鉴定动物双歧杆菌方面对实验设备及试剂的高要求,具有成本低、特异性强的特点,有较为广阔的市场开发前景。
    Abstract: In this study, in order to overcome the shortcomings of the current methods used to identify Bifidobacterium animalis, such as long time, complicated operation and low adaptability of experimental environment, specific primer probes were designed based on ERIC-PCR technology to identify and detect B.animalis.Based on the genomic DNA of B.animalis HP-B1124, the ERIC-PCR reaction conditions of B.animalis HP-B1124 were optimized, and the ERIC-PCR fragments were obtained one by one and sequenced.Two pairs of specific primer probes were designed.The accuracy, specificity, limitation and universality of the two pairs of primer probes were evaluated, and the two pairs of specific primer probes were used for testing the products containing B.animalis in the commercially published formula.The two pairs of specific primer probes designed in this study could be used for identified strains of B.animalis more simply, quickly and targeted.This method has optimized the current relatively traditional methods of pure culture and plate counting identification of B.animalis, and has solved the high requirements of SNP genotyping technology and real-time fluorescence quantitative PCR method for experimental equipment and reagents in the identification of B.animalis to a certain extent.It has the characteristics of low cost, high specificity and earn a broad market development prospect.
  • [1] Sharples GJ, Lloyd RG. A novel repeated DNA sequence located in the intergenic regions of bacterial chromosomes[J]. Nucleic Acids Res, 1990, 18(22): 6503-6508.
    [2] Cao YF, Zhao LP. Computational analysis of the distribution of ERIC (IRU) in different bacterial genomes[J]. J Shanxi Univ Nat Sci Ed (山西大学学报 自然科学版), 2002, 25(4): 354-357.
    [3] Zhang ZC, Yang ES, Guo KK, et al. ERIC-PCR fingerprint analysis of intestinal flora of diarrhea piglets[J]. J Northwest A F Univ Nat Sci Ed (西北农林科技大学学报 自然科学版), 2019, 47(1): 31-38.
    [4] Hematzadeh A, Haghkhah M. Biotyping of isolates of Pseudomonas aeruginosa isolated from human infections by RAPD and ERIC-PCR[J]. Heliyon, 2021, 7(9): e07967.
    [5] Shrivastava UP. Molecular diversity assessment of PGPR using enterobacterial repetitive intergeneric consensus (ERIC) PCR[J]. Acad Voices, 2018, 7: 64-70.
    [6] Versalovic J, Koeuth T, Lupski JR. Distribution of repetitive DNA sequences in eubacteria and application to fingerprinting of bacterial genomes[J]. Nucleic Acids Res, 1991, 19(24): 6823-6831.
    [7] Tian M, Feng Z, Qu RD, et al. Identification of Chryseobacterium proteolyticum from different regions using ERIC-PCR and automated ribotyping[J]. Sci Technol Food Ind (食品工业科技), 2020, 41(1): 105-111, 118.
    [8] Chen YC, Cao YF, Zhao LP. Non-random nature of genomic DNA amplification of E.coli K-12 MG1655 via ERIC-PCR[J]. Microbiol China (微生物学通报), 2002, 29(6): 28-32.
    [9] Li P, Li JX, Li YF, et al. Analysis of ERIC-PCR fingerprints of Haemophilus parasuis field isolates from south-east of China[J]. Chin J Vet Sci (中国兽医学报), 2009, 29(12): 1566-1570.
    [10] Chen JC, Zhang JY, Zhan L, et al. Prevalence and antimicrobial-resistant characterization of Bacillus cereus isolated from ready-to-eat rice products in Eastern China[J]. Front Microbiol, 2022, 13: 964823.
    [11] Khoshbayan A, Golmoradi Zadeh R, Taati Moghadam M, et al. Molecular determination of O25b/ST131 clone type among extended spectrum β-lactamases production Escherichia coli recovering from urinary tract infection isolates[J]. Ann Clin Microbiol Antimicrob, 2022, 21(1): 35.
    [12] Zhang TF, Wang J, Yao Z, et al. Effect and mechanism of Bifidobacterium animalis B94 in the prevention and treatment of liver injury in rats[J]. Front Cell Infect Microbiol, 2022, 12: 914684.
    [13] Do MH, Oh MJ, Lee HB, et al. Bifidobacterium animalis ssp. lactis MG741 reduces body weight and ameliorates nonalcoholic fatty liver disease via improving the gut permeability and amelioration of inflammatory cytokines[J]. Nutrients, 2022, 14(9): 1965.
    [14] Lv XB, He LP, Zhang RJ, et al. Research progress in physiological functions of Bifidobacterium[J]. Sci Technol Food Ind (食品工业科技), 2013, 34(16): 353-358.
    [15] Liu H, Zhang KG, Liu P, et al. Improvement effect of Bifidobacterium animalis subsp. lactis MH-02 in patients receiving resection of colorectal polyps: a randomized, double-blind, placebo-controlled trial[J]. Front Immunol, 2022, 13: 940500.
    [16] Liu K, Kong XJ. Altered salivary microbiota following Bifidobacterium animalis subsp. Lactis BL-11 supplementation are associated with anthropometric growth and social behavior severity in individuals with prader-willi syndrome[J]. Probiotics Antimicrob Proteins, 2022, 14(4): 699-711.
    [17] Li L, Li JN, Yu WY. Brief introduction of the method of bacterial classification and identification[J]. J Anhui Agric Sci (安徽农业科学), 2004, 32(3): 549-551.
    [18] Deng MK, Sun Y, Han WQ. Methods of identification of bacteria[J]. Prog Biomed Eng (生物医学工程学进展), 2014, 35(2): 84-88.
    [19] Kim Y, Oh S, Park S, et al. Lactobacillus acidophilus reduces expression of enterohemorrhagic Escherichia coli O157: H7 virulence factors by inhibiting autoinducer-2-like activity[J]. Food Control, 2008, 19(11): 1042-1050.
    [20] Lomonaco S, Furumoto EJ, Loquasto JR, et al. Development of a rapid SNP-typing assay to differentiate Bifidobacterium animalis ssp. lactis strains used in probiotic-supplemented dairy products[J]. J Dairy Sci, 2015, 98(2): 804-812.
    [21] Zhao YP, Guo YW, Liu LF, et al. Determination of animal Bifidobacterium BB-12 in yogurt products by TaqMan-MGB fluorescent quantitative polymerase chain reaction[J]. J Food Saf Qual(食品安全质量检测学报), 2021, 12(24): 9362-9370.
    [22] Pang J, Liu ZY, Hao M, et al. Comparison and optimization of methods for genomic DNA extraction from Gram positive bacteria[J]. Microbiol China (微生物学通报), 2015, 42(12): 2482-2486.
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出版历程
  • 收稿日期:  2022-09-11
  • 修回日期:  2023-03-26
  • 刊出日期:  2023-04-24

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