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首页> 外文期刊>Biotechnology & Biotechnological Equipment >EVOLUTION OF CRISPR1 AND CRISPR3 IN SPONTANEOUS PHAGE-RESISTANT MUTANTS OF STREPTOCOCCUS THERMOPHILUS STRAIN LBB.A
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EVOLUTION OF CRISPR1 AND CRISPR3 IN SPONTANEOUS PHAGE-RESISTANT MUTANTS OF STREPTOCOCCUS THERMOPHILUS STRAIN LBB.A

机译：CRISPR1和CRISPR3在嗜热链球菌LBB.A自发噬菌体突变中的进化

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摘要

Streptococcus thermophilus LBB.A is a component of industrial starters for traditional Bulgarian yogurt. Eight genetically distinguishable cos-type phage cultures: phi A1, phi A2, phi A5, phi A6, phi DmA, phi SfA, phi PtA, and phi A7, were found to infect strain LBB.A. Bacteriophage-Resistant Mutants (BIMs) of strain LBB.A were isolated and their Clustered Regularly Interspaced Short Palindrome Repeats (CRISPR1 and CRISPR3) were amplified by PCR and compared to the parent strain. Challenging S. thermophilus LBB.A with phi A1 resulted in the isolation of "first order" BIM AR(3)(A1), which was resistant to six of the phages, but remained sensitive to phi PtA and phi A7. In AR(3)(A1) the size of the CRISPR1 and CRISPR3 loci increased with approximately 66 bp each, compared to the parent strain, corresponding to the insertion of one additional repeat unit. Next, AR(3)(A1) was challenged with phi A7 to obtain "second order" BIM AR(3-3)(AIA7), which was resistant to all of the eight phages. Strain AR(3-3)(AIA7) was found to have acquired only another 65 bp in its CRISPR1 locus with no change in CRISPR3. The newly acquired sequences in CRISPR1 and CRISPR3 of AR(3)(AI) and AR(3-3)(AIA7) showed similarity to sequences in already published S. thermophilus phage genomes. The phage-resistant mutant AR(3-3)(AIA7) acidified milk at a rate comparable to the parent strain LBB.A and, consequently, AR(3-3)(AIA7) was recommended as a potential replacement of the sensitive parent strain in yogurt production. The difference in the size of the amplified region of CRISPR1 and CRISPR3 can be used as a genetic marker to differentiate between BIMs and their phage sensitive parent strain.
机译：嗜热链球菌LBB.A是传统保加利亚酸奶的工业发酵剂的组成部分。发现八个遗传上可区分的cos型噬菌体培养物：phi A1，phi A2，phi A5，phi A6，phi DmA，phi SfA，phi PtA和phi A7，感染了菌株LBB.A.分离了菌株LBB.A的抗噬菌体突变体（BIM），并通过PCR扩增了它们的成簇的规则间隔的短回文重复序列（CRISPR1和CRISPR3），并与亲本菌株进行了比较。用phi A1挑战嗜热链球菌LBB.A导致分离出“一级” BIM AR（3）（A1），它对六个噬菌体具有抗性，但仍对phi PtA和phi A7敏感。在AR（3）（A1）中，与亲本菌株相比，CRISPR1和CRISPR3基因座的大小分别增加了约66 bp，相当于插入了一个额外的重复单元。接下来，用phi A7攻击AR（3）（A1），以获得对所有八个噬菌体均具有抗性的“二阶” BIM AR（3-3）（AIA7）。发现菌株AR（3-3）（AIA7）在其CRISPR1基因座中仅获得了另外65 bp，而CRISPR3没有变化。 AR（3）（AI）和AR（3-3）（AIA7）的CRISPR1和CRISPR3中新获得的序列与已发表的嗜热链球菌噬菌体基因组中的序列相似。抗噬菌体的突变体AR（3-3）（AIA7）以与亲本菌株LBB.A相当的速率酸化了牛奶，因此建议将AR（3-3）（AIA7）用作敏感亲本的潜在替代品酸奶生产中的应变。 CRISPR1和CRISPR3扩增区域大小的差异可以用作遗传标记，以区分BIM及其噬菌体敏感亲本菌株。

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