CN110106113B - 一株高加索酸奶乳杆菌msr101及其应用 - Google Patents
一株高加索酸奶乳杆菌msr101及其应用 Download PDFInfo
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- CN110106113B CN110106113B CN201910388000.6A CN201910388000A CN110106113B CN 110106113 B CN110106113 B CN 110106113B CN 201910388000 A CN201910388000 A CN 201910388000A CN 110106113 B CN110106113 B CN 110106113B
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- kefiri
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Abstract
本发明涉及一株高加索酸奶乳杆菌Lactobacillus kefiri MSR101,属于乳酸菌技术领域。本发明提供的高加索酸奶乳杆菌Lactobacillus kefiri MSR101,其保藏编号为CGMCC No.17506。本发明所述高加索酸奶乳杆菌具有耐酸性,耐胆汁盐性,耐酚性,抗生素耐性,抗氧化活性,细胞表面疏水性,对肠上皮细胞的粘附能力和降低胆固醇的功能。
Description
技术领域
本发明涉及乳酸杆菌技术领域,具体涉及一株高加索酸奶乳杆菌Lactobacilluskefiri MSR101及其应用。
背景技术
益生菌微生物是“活微生物”,可以为宿主带来各种健康益处,包括免疫系统的调节,胃肠道微生物群的重排和抑制有害微生物的生长(Maleki Kakelar等,2019)。乳酸菌是最重要的益生菌,并且与人体消化系统具有相容性,因为它们具有对低pH值和高胆汁盐条件的天然抗性(Shehata等,2016,Riaz Rajoka等,2018)。
近10年来,益生菌已成为乳酸菌领域的研究热点,其中包括乳酸杆菌属和双歧杆菌属。(Bao等,2010)。最近,对从具有各种健康促进作用的中国传统乳制品中分离的乳杆菌菌株的消耗量有了新的认识。为了提供给宿主健康所需的条件,分离的乳杆菌菌株必须具备通过胃肠道的物理和化学屏障的能力(Ramos等,2018)。此外,作为益生菌,在预期产品的生产和储存期间,乳杆菌菌株必须具有存活足够数量的能力。
发明内容
本发明的目的在于提供一株高加索酸奶乳杆菌Lactobacillus kefiri MSR101及其应用。本发明所述乳杆菌具有耐酸性,耐胆汁盐性,耐酚性,抗生素耐性,抗氧化活性,细胞表面疏水性,对肠上皮细胞的粘附能力和降低胆固醇的功能。
本发明提供了一株高加索酸奶乳杆菌Lactobacillus kefiri MSR101,其特征在于,保藏编号为CGMCCNo.17506。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在抗高胆汁盐中的应用。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在苯酚耐受中的应用。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在抗氧化中的应用。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在降低胆固醇浓度中的应用。
本发明提供了一株高加索酸奶乳杆菌Lactobacillus kefiri MSR101。本发明所述乳杆菌具有耐酸性,耐胆汁盐性,耐酚性,抗生素耐性,抗氧化活性,细胞表面疏水性,对肠上皮细胞的粘附能力和降低胆固醇的功能。试验结果表明,高加索酸奶乳杆菌Lactobacillus kefiri MSR101菌株培养48h后处于稳定期且EPS产量最高达753mg/L,高加索酸奶乳杆菌Lactobacillus kefiri MSR101菌株对二甲苯显示出较高的疏水性(70%),对肠上皮细胞HT-29的最高粘附率为34.4%,对超氧化物阴离子自由基的清除率为65.5%,对DPPH自由基清除率为43.3%,胆固醇去除率为26.2%。
附图说明
图1为本发明提供的系统发育树分析结果图;
图2为本发明提供的抗生素耐药性结果图;
图3为本发明提供的胞外多糖产量结果图。
生物保藏说明
高加索酸奶乳杆菌Lactobacillus kefiri MSR101。本菌种保藏在中国微生物菌种保藏管理委员会普通微生物中心,地址为北京市朝阳区北辰西路1号院3号,中国科学院微生物研究所,保藏编号为CGMCC No.17506,保藏时间为2019年4月1日。
具体实施方式
本发明提供了一株高加索酸奶乳杆菌Lactobacillus kefiri MSR101,保藏编号为CGMCCNo.17506。
高加索酸奶乳杆菌Lactobacillus kefiri MSR101是一种厌氧、革兰氏阳性菌,杆状细菌。
本发明所述高加索酸奶乳杆菌Lactobacillus kefiri MSR101是革兰氏阳性,过氧化氢酶阴性,杆状和嗜温的。其16s rRNA基因序列如SEQ ID NO.1所示:
GCTTGGCGTCGTGCTATACATGCAAGTCGAACGCGTTTCCGTTATT GATTTTAGAGTGTTGCATTTGAATGATTTAACACGAAACGAGTGGCGAACTGGTGAGTAACACGTGGGTACCTGCCCTTGAAGTAGGGGATAACACTTGGAAACAGGTGCTAATACCGTATAACAACCAAAACCACATGGTTTTGGTTTAAAAGATGGCTTCGGCTATCACTTTAGGATGGACCCGGGCGTATTAGCTTGTTGGTAAGGTAATGGCCTACCAAGGCAATGATACGTAGCCGACCTAGAGGGTAATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGGTAGGGAATCTTCCACAATGGACGAAAGTCTGATGGAGCAACGCCGCGTGAGTGATGAAGGGTTTCGGCTCGTAAAACTCTGTTGTTGGAGAAGAACAGGTGTCAGAGTAACTGTTGACATCTTGACGGTATCCAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTCTTAGGTCTGATGTGAAAGCCTTCGGCTTAACCGGAGAAGTGCATCGGAAACCAGGAGACTTGAGTGCAGAAGAGGACAGTGGAACTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCATGGGTAGCGAACAGGATTAGATACCCTGGTAGTCCATGCCGTAAACGATGAGTGCTAAGTGTTGGAGGGTTTCCGCCCTTCAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGAGCATGTGTTTAATTCGATGCTACGCGAAGACCTTACCAGGTCTTGACATCTTTCTCCTATTTCTGTCACCTTAGACGGCTGGTCCCCGAAGGTTA。
所述序列显示其与NCBI数据库中可获得的已知物种的序列相似99%,并且序列以登录号MK491609保藏在Gene Bank中。在抗生素敏感性,肠道条件下耐受性(低pH,胆汁盐耐受性和0.2%至0.4%酚抗性),疏水性,抗氧化能力,降低胆固醇和对HT-29的粘附等方面有有益效果。所述高加索酸奶乳杆菌Lactobacillus kefiri MSR101对青霉素,氨苄青霉素,链霉素和四环素具有抗性。此外,它还表现出对HT-29细胞的粘附性(34.3%)和抗氧化活性(67%)。在胃肠道条件下显示出高的存活率(>80%)。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在抗高胆汁盐中的应用。本发明菌株在高胆汁盐条件下(0.3%、0.5%和1%胆汁(w/v)).菌株生长状态良好。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在苯酚耐受中的应用。本发明所述菌株在苯酚浓度为0.2%~0.4%下具有良好的存活能力。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在抗氧化中的应用。本发明所述菌株具有抗氧化活性(67%)。
本发明还提供了上述技术方案所述高加索酸奶乳杆菌Lactobacillus kefiriMSR101在降低胆固醇浓度中的应用。本发明所述菌株降低胆固醇能力为26.2±0.6%。
下面结合具体实施例对本发明所述的一株高加索酸奶乳杆菌Lactobacilluskefiri MSR101及其应用做进一步详细的介绍,本发明的技术方案包括但不限于以下实施例。
实施例1
采样
实验样品来自中国青海省西藏自治区的中国传统乳制品,共有20份,分别有牦牛奶(4)、马乳酒(5)、奶酪(4)和克菲尔谷物(7)。将样品收集在无菌管中并保存在-15℃的迷你冷冻机中。实验室收到样品后立即进行了试验。
乳酸杆菌的分离和鉴定
将样品在磷酸盐缓冲液(PBS,pH 7.2)中连续稀释,并将每批稀释液的小等分试样(50μl)涂布在补充有0.05%L-半胱氨酸(MRSc)的de Man-Rogosa-Sharpe(Merck,China)琼脂平板上。将平板在37℃下厌氧培养72h。选择具有不同形态的菌落并在新制备的MRSc琼脂平板上重新划线数代,分离纯化单个菌落。所有分离株均经革兰染色和过氧化氢酶试验证实为乳酸菌,革兰阳性和过氧化氢酶阴性的菌株均为乳酸菌。通过对16S rDNA基因测序,进行乳杆菌分离物的鉴定。使用基因组DNA纯化试剂盒(TransGen Biotech Co.,Ltd.,Beijing,China),按照使用说明提取总基因组DNA。用于扩增16S rDNA序列的引物是正向5“-AGAGTTTGATCCTGGCTC AG-3”(SEQ ID NO.2)和反向5“-CCGTCAATTCCTTTGAGTTT-3”(SEQID NO.3)。在Techne-TC 512热循环仪(英国)中,在以下条件下扩增片段:95℃,1min;95℃,30s,30个循环;55℃,30秒,最后72℃,5min。将扩增的片段在琼脂糖凝胶上筛选,并由中国深圳的广州IGE生物技术有限公司测序。通过BLAST程序(https://blast.ncbi.nlm.nih.gov/Blast.cgi)测试所有获得的序列。序列保存在基因库中。通过ClustalW2(http://www.ebi.ac.uk/Tool/mas/clustalw2/)进行序列比对,通过邻接(Tamura等,2004)和最大复合似然法构建系统发育树。使用Mega 6.0软件(http://megasoftware.net/)(Saitou和Nei,1987)。结果表明,克菲尔活菌元是乳酸杆菌的来源之一。
系统发育树分析结果如图1所示,系统发育树是在16S rRNA序列的基础上构建的。基因组概率用1000个重复数确定,并以百分比值表示。实心圆表示菌株来自NCBI,空心圆表示分离的乳酸杆菌菌株用于发育树构建。
实施例2
在低pH值和高胆汁盐条件下的生存状况
按照所(Lee等人,2011)描述的方法评估乳酸杆菌在pH2.0,pH2.5,pH3.0和pH6.5(对照组)下的存活能力。同时,使用含有0.3%,0.5%和1%胆汁(w/v)的MRSc肉汤培养基,按照(Sabir等人,2010b)方法测定在酸性条件下存活3小时的乳酸杆菌的胆汁盐耐受性。没有胆汁盐的MRSc肉汤培养基用作对照。根据公式(1),通过MRSc琼脂平板上的计数评估乳酸杆菌对酸性条件以及胆汁盐条件的抗性。
增长率(%)=(N1/N0)×100 公式(1)
其中,N1是处理后MRSc肉汤中活细菌细胞的总数,N0是处理前MRSc肉汤中活细菌细胞的总数。结果表明,在低pH值(pH2.0、pH2.5pH3.0)和高胆汁盐条件下(0.3%、0.5%和1%胆汁(w/v)).菌株MSR101生长状态良好。
实施例3
苯酚耐受性
苯酚耐受性测定通过先前报道(Shehata等人,2016)的方法进行,对其稍作修改。将1%乳酸菌份分离物培养24h后接种于添加了0.2%苯酚和0.4%苯酚的MRsc发酵液中。将不含苯酚的MRSc肉汤培养基用作对照。在37℃下,温育24h后,按公式测定吸光度(A)OD630nm来评价酚抗性。
苯酚耐受性(%)=(A1/A0)×100
其中,A1是处理后培养物的吸光度,A0是处理前培养物的吸光度。结果表明,菌株MSR101在苯酚浓度为0.2%~0.4%下具有良好的存活能力。
实施例4
溶血活性
乳酸杆菌分离物以条纹状分布在有5%羊血的哥伦比亚血培养基(Sigma,中国)的表面上,并将平板在37℃下培养72h。温育后,检查平板的溶血活性。
结果表明,MSR101菌株无溶血活性,因此在溶血活性方面是安全的。
实施例5
抗生素耐药性
通过之前报道过的(Riaz Rajoka等,2017a)琼脂重叠扩散法评估乳酸杆菌分离物的抗生素耐药性。检测了所有乳酸杆菌分离物对10种抗生素的抗性,包括青霉素,红霉素,氨苄青霉素,链霉素,四环素,万古霉素,庆大霉素,卡那霉素,氯霉素和克林霉素。方法是将新鲜制备的MRSc琼脂平板用50μl的活性乳酸杆菌培养物覆盖,并将平板在4℃孵育1h,将抗生素盘置于平板上,并将平板在37℃下温育24h后,测量抑制区的直径。抗生素耐药性结果如图2和表1所示,所述高加索酸奶乳杆菌Lactobacillus kefiri MSR101对青霉素,氨苄青霉素,链霉素和四环素具有抗性。
表1抗生素耐药性
*(S)=易感染and(R)=免疫
实施例6
胞外多糖的生产
乳酸杆菌培养物在新鲜制备的100ml并补充有3%(w/v)葡萄糖的MRSc肉汤的烧瓶中生长,并在37℃下孵育72h。通过离心(7000×g,10min)除去细菌细胞,并将两体积的预冷乙醇加入到一体积的上清液中,用于胞外多糖(EPS)沉淀。通过在4℃下离心(10000×g,35min)回收EPS沉淀,透析(6000Da至8000Da)48h,然后冻干。使用葡萄糖作为标准液,用苯酚硫酸法(Nikolic等人,2012)测量糖的总量。目前,生产EPS的L.kefiri MSR101菌株最初是从青海省西藏自治区开菲尔谷物中分离得到的,MSR101菌株显示出粘稠的外观,这表明它是一株EPS生产菌株。L.kefiri MSR101菌株培养48h后处于稳定期且EPS量达到最大值753mg/L,随后进入衰亡期。结果表明,MSR101菌株在培养的32h内生长迅速,同时培养基pH值快速下降,而48h后,该菌株生长速度下降,培养基pH值略有下降,在48h时pH值约为3.1±0.5。
实施例7
细胞表面疏水性
通过微生物对烃类化合物的粘附试验,对乳酸菌的细胞表面疏水性进行了研究(Kotzamanidis等人,2010)。简言之,通过离心(8000×g,5min)收获静止期细胞,用PBS(pH7.2)洗涤两次,最后重悬浮于PBS(pH7.2)中,在630nm处达到0.6±0.02的OD(A0)。将1ml二甲苯与1ml细胞悬浮液混合,并在室温下静置30min以形成两相体系。小心地除去水相,测量其在630nm处的吸光度(A1)。通过下面的公式测量细胞表面疏水性(%):
细胞表面疏水性(%)=(1-A1/A0)×100
Lactobacillus kefiri MSR101菌株对二甲苯显示出较高的疏水性(70%);二甲苯是测量微生物细胞疏水性的标准碳氢化合物。
实施例8
抗氧化活性
制备完整细胞
将过夜的乳酸杆菌培养物离心(7000×g,4℃下15min),用PBS(pH7.2)洗涤细胞三次,最后重悬于PBS(pH7.2)中,终浓度为1×106cfu/ml,进行抗氧化剂分析。
超氧阴离子清除试验
采用改进的邻苯三酚自氧化法(Re等人,2014年)进行超氧阴离子清除实验。即将PBS(pH7.2)中最终浓度为1×106cfu/ml的乳酸杆菌细胞悬浮液(100μl)、900μl水与2mlTric-HCl缓冲液(pH8.1)快速混合。使用无菌蒸馏水代替乳酸杆菌细胞悬浮液作为对照。加入50μmol/ml邻苯三酚溶液,孵育10min后测定对照组和样品在330nm处的吸光度,评价其自氧化作用。通过以下公式计算测试的乳酸杆菌的超氧阴离子清除能力:
清除能力(%)=(ΔA0-ΔA)×100/ΔA0
其中ΔA0和ΔA分别是加入样品和去离子水之前和之后的连苯三酚的自氧化速率。超氧化物阴离子自由基的清除率为65.5%。
DPPH自由基清除活性
通过先前报道的方法(Son等,2018)测定乳酸杆菌的DPPH自由基清除能力。即将三(3)ml乳酸杆菌细胞悬浮液和3ml 0.5mM DPPH自由基的乙醇溶液混合,并在室温下黑暗中温育30min。温育后,将反应混合物离心(10000×g,4℃下20min),测量上清液在517nm处的吸光度。通过下式计算DPPH清除能力:
清除能力(%)=(1-ODsample/ODcontrol)×100
其中ODSample和ODControl分别是样品和蒸馏水的吸光度,与DPPH溶液混合。Lactobacillus kefiri MSR101对DPPH自由基清除率为43.3%。
实施例9
降低胆固醇能力
分离的乳酸菌对胆固醇的吸收能力是按照对之前报道的方法(Wang et al.,2012)进行修改后测定的。简言之,将每种乳酸杆菌分离物(1%v/v)接种到10ml新鲜制备的MRSc肉汤培养基中,该肉汤含有0.2%硫代乙酸钠(Sigma,中国),0.3%牛胆汁(Sigma,中国)和100μl/ml水溶性胆固醇(Sigma,中国),将每个试管在37℃下孵育24h。通过离心(10,000×g,4℃下35min)除去细胞。用测量胆固醇的邻苯二甲醛方法(Rudel和Morris,1973)来测定使用过的肉汤和未接种的无菌肉汤中的胆固醇量。菌株Lactobacillus kefiriMSR101的胆固醇去除率为26.2%。
实施例10
粘附测定
使用添加有2mM L-谷氨酰胺,10%热灭活的胎牛血清,100μg链霉素/ml,1%非必需氨基酸和100Ul的Dulbecco改良Eagle培养基(DMEM),在细胞培养瓶中培养HT-29细胞。随后将HT-29细胞以2.5×105细胞/孔的浓度接种到24孔培养板中,培养3天,每天更换培养基。将细胞在37℃,5%CO2气氛中温育。将乳杆菌分离物的过夜培养物离心,用PBS(0.1M,pH7.2)洗涤两次,并在相同的缓冲液中重悬至适当浓度的稀释液(吸光度OD6300.2,约2×108CFU/ml)。然后将细菌细胞加入到每个细胞孔中,37℃孵育4h,孵育后用PBS(0.1M,pH7.2)冲洗细胞,0.1%TritonX-100溶液裂解。细胞裂解液经连续稀释后涂于MRSc琼脂平板上。37℃孵育3天。计算细菌粘附率:
黏附率%=(附着菌/总添加菌)×100
菌株Lactobacillus kefiri MSR101在HT-29细胞中的最高的粘附率为34.4%。
实施例11
统计分析
数值以平均值和标准偏差给出,平行三次测定。Tukey多重比较检验在所有试验中均有显著性差异(p<0.05),有统计学意义。
技术效果:
乳酸杆菌kefiri MSR101及其益生菌潜力
从中国青海省西藏自治区收集的克非尔产品样品中分离出乳酸杆菌MSR101。它是革兰氏阳性,过氧化氢酶阴性,杆状和嗜温的。基于16s rRNA基因序列(GCTTGGCGTCGTGCTATACATGCAAGTCGAACGCGTTTCCGTTATTGATTTTAGAGTGTTGCATTTGAATGATTTAACACGAAACGAGTGGCGAACTGGTGAGTAACACGTGGGTACCTGCCCTTGAAGTAGGGGATAACACTTGGAAACAGGTGCTAATACCGTATAACAACCAAAACCACATGGTTTTGGTTTAAAAGATGGCTTCGGCTATCACTTTAGGATGGACCCGGGCGTATTAGCTTGTTGGTAAGGTAATGGCCTACCAAGGCAATGATACGTAGCCGACCTAGAGGGTAATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGGTAGGGAATCTTCCACAATGGACGAAAGTCTGATGGAGCAACGCCGCGTGAGTGATGAAGGGTTTCGGCTCGTAAAACTCTGTTGTTGGAGAAGAACAGGTGTCAGAGTAACTGTTGACATCTTGACGGTATCCAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTCTTAGGTCTGATGTGAAAGCCTTCGGCTTAACCGGAGAAGTGCATCGGAAACCAGGAGACTTGAGTGCAGAAGAGGACAGTGGAACTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCATGGGTAGCGAACAGGATTAGATACCCTGGTAGTCCATGCCGTAAACGATGAGTGCTAAGTGTTGGAGGGTTTCCGCCCTTCAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGAGCATGTGTTTAATTCGATGCTACGCGAAGACCTTACCAGGTCTTGACATCTTTCTCCTATTTCTGTCACCTTAGACGGCTGGTCCCCGAAGGTTA)鉴定分离物,并且获得的序列显示其与NCBI数据库中可获得的已知物种的序列相似99%,并且序列以登录号MK491609保藏在Gene Bank中。在抗生素敏感性,肠道条件下耐受性(低pH,胆汁盐耐受性和0.2%至0.4%酚抗性),疏水性,抗氧化能力,降低胆固醇和对HT-29的粘附等方面测试益生菌性质。本发明菌株对青霉素,氨苄青霉素,链霉素和四环素具有抗性。此外,它还表现出对HT-29细胞的粘附性(34.3%)和抗氧化活性(67%)。在胃肠道条件下显示出高的存活率(>80%),表明它们在益生菌应用方面有潜力(如表2所示)。
表2益生菌潜力
以上所述仅是本发明的优选实施方式,应当指出,对于本技术领域的普通技术人员来说,在不脱离本发明原理的前提下,还可以做出若干改进和润饰,这些改进和润饰也应视为本发明的保护范围。
序列表
<110> 深圳大学
<120> 一株高加索酸奶乳杆菌MSR101及其应用
<160> 3
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1032
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
gcttggcgtc gtgctataca tgcaagtcga acgcgtttcc gttattgatt ttagagtgtt 60
gcatttgaat gatttaacac gaaacgagtg gcgaactggt gagtaacacg tgggtacctg 120
cccttgaagt aggggataac acttggaaac aggtgctaat accgtataac aaccaaaacc 180
acatggtttt ggtttaaaag atggcttcgg ctatcacttt aggatggacc cgggcgtatt 240
agcttgttgg taaggtaatg gcctaccaag gcaatgatac gtagccgacc tagagggtaa 300
tcggccacat tgggactgag acacggccca aactcctacg ggaggcaggt agggaatctt 360
ccacaatgga cgaaagtctg atggagcaac gccgcgtgag tgatgaaggg tttcggctcg 420
taaaactctg ttgttggaga agaacaggtg tcagagtaac tgttgacatc ttgacggtat 480
ccaaccagaa agccacggct aactacgtgc cagcagccgc ggtaatacgt aggtggcaag 540
cgttgtccgg atttattggg cgtaaagcga gcgcaggcgg ttcttaggtc tgatgtgaaa 600
gccttcggct taaccggaga agtgcatcgg aaaccaggag acttgagtgc agaagaggac 660
agtggaactc catgtgtagc ggtgaaatgc gtagatatat ggaagaacac cagtggcgaa 720
ggcggctgtc tggtctgtaa ctgacgctga ggctcgaaag catgggtagc gaacaggatt 780
agataccctg gtagtccatg ccgtaaacga tgagtgctaa gtgttggagg gtttccgccc 840
ttcagtgctg cagctaacgc attaagcact ccgcctgggg agtacgaccg caaggttgaa 900
actcaaagga attgacgggg gcccgcacaa gcggtgagca tgtgtttaat tcgatgctac 960
gcgaagacct taccaggtct tgacatcttt ctcctatttc tgtcacctta gacggctggt 1020
ccccgaaggt ta 1032
<210> 2
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
agagtttgat cctggctcag 20
<210> 3
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 3
ccgtcaattc ctttgagttt 20
Claims (2)
1.一株高加索酸奶乳杆菌Lactobacillus kefiri MSR101,其特征在于,保藏编号为CGMCC No.17506。
2.权利要求1所述高加索酸奶乳杆菌Lactobacillus kefiri MSR101在苯酚耐受中的应用。
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