title: ÐÐ·ÑÑÐµÐ½Ð¸Ðµ ÑÐ¾Ð»Ð¸ Ð±ÐµÐ»ÐºÐ¾Ð² ÑÐµÐ¿Ð°ÑÐ°ÑÐ¸Ð¸ Ð² Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð¾Ð¹ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ð¸ Ð² ÑÐ¸ÑÑÐµÐ¼Ðµ CRISPR-Cas I-E Escherichia coli
title: Ð²ÑÐ¿ÑÑÐºÐ½Ð°Ñ ÐºÐ²Ð°Ð»Ð¸ÑÐ¸ÐºÐ°ÑÐ¸Ð¾Ð½Ð½Ð°Ñ ÑÐ°Ð±Ð¾ÑÐ° Ð±Ð°ÐºÐ°Ð»Ð°Ð²ÑÐ°
titleType: Subtitle
title: Study of the role of DNA repair proteins in primed adaptation in the type I-E CRISPR-Cas system of Escherichia coli
Creators: name: Ð¦Ð¾Ð¹, Ð®Ð»Ð¸Ñ
Publisher: name: Ð¡Ð°Ð½ÐºÑ-ÐÐµÑÐµÑÐ±ÑÑÐ³ÑÐºÐ¸Ð¹ Ð¿Ð¾Ð»Ð¸ÑÐµÑÐ½Ð¸ÑÐµÑÐºÐ¸Ð¹ ÑÐ½Ð¸Ð²ÐµÑÑÐ¸ÑÐµÑ ÐÐµÑÑÐ° ÐÐµÐ»Ð¸ÐºÐ¾Ð³Ð¾
Year: 2022
Subjects: CRISPR-Cas
Escherichia coli
Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð°Ñ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ñ
Ð¿ÑÐµÑÐ¿ÐµÐ¹ÑÐµÑÑ
primed adaptation
prespacers

description: ÐÐ°Ð´Ð°ÑÐ¸ Ð¸ÑÑÐ»ÐµÐ´Ð¾Ð²Ð°Ð½Ð¸Ñ: - Ð¿Ð¾Ð»ÑÑÐ¸ÑÑ ÑÑÐ°Ð¼Ð¼Ñ Ñ Ð´ÐµÐ»ÐµÑÐ¸ÑÐ¼Ð¸ âxseA, ârecJ âxseA, ârecQ, ârecB ârecQ; - Ð¸Ð·ÑÑÐ¸ÑÑ ÑÐ¿Ð¾ÑÐ¾Ð±Ð½Ð¾ÑÑÑ Ð½ÑÐºÐ»ÐµÐ°Ð·Ñ ExoVII Ð·Ð°Ð¼ÐµÑÐ°ÑÑ Ð°ÐºÑÐ¸Ð²Ð½Ð¾ÑÑÑ RecJ Ð² Ð¿ÑÐ¾ÑÐµÑÑÐµ Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð¾Ð¹ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ð¸ Ð½Ð° ÑÐ¾Ð½Ðµ Ð´ÐµÐ»ÐµÑÐ¸Ð¸ Ð³ÐµÐ½Ð° recJ; - Ð¸Ð·ÑÑÐ¸ÑÑ ÑÐ¿Ð¾ÑÐ¾Ð±Ð½Ð¾ÑÑÑ ÑÐµÐ»Ð¸ÐºÐ°Ð·Ñ RecQ Ð·Ð°Ð¼ÐµÑÐ°ÑÑ ÑÐµÐ»Ð¸ÐºÐ°Ð·Ð½ÑÑ Ð°ÐºÑÐ¸Ð²Ð½Ð¾ÑÑÑ RecBCD Ð² Ð¿ÑÐ¾ÑÐµÑÑÐµ Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð¾Ð¹ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ð¸ Ð½Ð° ÑÐ¾Ð½Ðµ Ð´ÐµÐ»ÐµÑÐ¸Ð¸ Ð³ÐµÐ½Ð° recB; - ÑÑÐ°Ð²Ð½Ð¸ÑÑ ÑÑÑÐµÐºÑÐ¸Ð²Ð½Ð¾ÑÑÑ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ð¸ Ð½Ð° ÑÐ¾Ð½Ðµ Ð¸Ð½Ð°ÐºÑÐ¸Ð²Ð°ÑÐ¸Ð¸ ÑÐµÐ»Ð¸ÐºÐ°Ð·Ð½Ð¾Ð¹ Ð°ÐºÑÐ¸Ð²Ð½Ð¾ÑÑÐ¸ RecD Ð² ÑÐµÐ·ÑÐ»ÑÑÐ°ÑÐµ ÑÐ¾ÑÐºÐ¾Ð²Ð¾Ð¹ Ð¼ÑÑÐ°ÑÐ¸Ð¸ Ð»Ð¸Ð±Ð¾ Ð² ÑÐµÐ·ÑÐ»ÑÑÐ°ÑÐµ Ð´ÐµÐ»ÐµÑÐ¸Ð¸ Ð³ÐµÐ½Ð° recD. ÐÐ±ÑÐµÐºÑÐ¾Ð¼ Ð¸ÑÑÐ»ÐµÐ´Ð¾Ð²Ð°Ð½Ð¸Ñ Ð´Ð°Ð½Ð½Ð¾Ð¹ ÑÐ°Ð±Ð¾ÑÑ ÑÐ²Ð»ÑÐµÑÑÑ ÑÑÐ°Ð¼Ð¼ Escherichia coli KD403 Ñ Ð°Ð²ÑÐ¾ÑÐ°ÑÐ³ÐµÑÐ¸ÑÑÑÑÐµÐ¹ ÑÐ¸ÑÑÐµÐ¼Ð¾Ð¹ CRISPR-Cas ÑÐ¸Ð¿Ð° I-E, ÐºÐ¾ÑÐ¾ÑÐ°Ñ Ð²ÐºÐ»ÑÑÐ°ÐµÑ Ð² ÑÐµÐ±Ñ Ð³ÐµÐ½Ñ cas Ð¿Ð¾Ð´ ÐºÐ¾Ð½ÑÑÐ¾Ð»ÐµÐ¼ Ð¸Ð½Ð´ÑÑÐ¸Ð±ÐµÐ»ÑÐ½ÑÑ Ð¿ÑÐ¾Ð¼Ð¾ÑÐ¾ÑÐ¾Ð² Ð¸ CRISPR-ÐºÐ°ÑÑÐµÑÑ Ñ ÐµÐ´Ð¸Ð½ÑÑÐ²ÐµÐ½Ð½ÑÐ¼ ÑÐ¿ÐµÐ¹ÑÐµÑÐ¾Ð¼, Ð¼Ð¸ÑÐµÐ½ÑÑ ÐºÐ¾ÑÐ¾ÑÐ¾Ð³Ð¾ ÑÐ²Ð»ÑÐµÑÑÑ Ð¿ÑÐ¾ÑÐ¾ÑÐ¿ÐµÐ¹ÑÐµÑ Ð² Ð³ÐµÐ½Ðµ yihN Ð² Ð³ÐµÐ½Ð¾Ð¼Ðµ ÐºÐ»ÐµÑÐºÐµ. ÐÑÐµÐ´Ð¼ÐµÑÐ¾Ð¼ Ð¸ÑÑÐ»ÐµÐ´Ð¾Ð²Ð°Ð½Ð¸Ñ ÑÐ²Ð»ÑÐµÑÑÑ Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð°Ñ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ñ Ð² ÑÐ¸ÑÑÐµÐ¼Ðµ I-E Ð¸ Ð²ÐºÐ»Ð°Ð´ ÑÐ°Ð·Ð»Ð¸ÑÐ½ÑÑ Ð³ÐµÐ½Ð¾Ð² ÑÐµÐ¿Ð°ÑÐ°ÑÐ¸Ð¸ Ð¸ ÑÐµÐºÐ¾Ð¼Ð±Ð¸Ð½Ð°ÑÐ¸Ð¸ Ð² ÑÑÑÐµÐºÑÐ¸Ð²Ð½Ð¾ÑÑÑ Ð´Ð°Ð½Ð½Ð¾Ð³Ð¾ Ð¿ÑÐ¾ÑÐµÑÑÐ°. Ð Ð°Ð±Ð¾ÑÐ° Ð±ÑÐ»Ð° Ð²ÑÐ¿Ð¾Ð»Ð½ÐµÐ½Ð° Ð½Ð° Ð±Ð°Ð·Ðµ Ð»Ð°Ð±Ð¾ÑÐ°ÑÐ¾ÑÐ¸Ð¸ ÐÐÐ Â«ÐÐ°Ð½Ð¾ÐÐ¸Ð¾Â» Ð¡Ð°Ð½ÐºÑ-ÐÐµÑÐµÑÐ±ÑÑÐ³ÑÐºÐ¾Ð³Ð¾ Ð³Ð¾ÑÑÐ´Ð°ÑÑÑÐ²ÐµÐ½Ð½Ð¾Ð³Ð¾ Ð¿Ð¾Ð»Ð¸ÑÐµÑÐ½Ð¸ÑÐµÑÐºÐ¾Ð³Ð¾ ÑÐ½Ð¸Ð²ÐµÑÑÐ¸ÑÐµÑÐ° ÐÐµÑÑÐ° ÐÐµÐ»Ð¸ÐºÐ¾Ð³Ð¾. Ð ÑÐ¾Ð´Ðµ Ð¸ÑÑÐ»ÐµÐ´Ð¾Ð²Ð°Ð½Ð¸Ñ Ð¿Ð¾Ð»ÑÑÐµÐ½Ñ Ð´Ð°Ð½Ð½ÑÐµ, Ð¿Ð¾Ð´ÑÐ²ÐµÑÐ¶Ð´Ð°ÑÑÐ¸Ðµ Ð·Ð°Ð¼ÐµÑÐµÐ½Ð¸Ðµ Ð² ÐºÐ»ÐµÑÐºÐ°Ñ ârecJ Ð½ÑÐºÐ»ÐµÐ°Ð·Ð½Ð¾Ð¹ Ð°ÐºÑÐ¸Ð²Ð½Ð¾ÑÑÐ¸ RecJ Ð½ÑÐºÐ»ÐµÐ°Ð·Ð¾Ð¹ ExoVII Ð² Ð¿ÑÐ¾ÑÐµÑÑÐµ Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð¾Ð¹ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ð¸. ÐÑÐ»Ð¾ Ð¿ÑÐ¾Ð´ÐµÐ¼Ð¾Ð½ÑÑÑÐ¸ÑÐ¾Ð²Ð°Ð½Ð¾, ÑÑÐ¾ Ð² ÐºÐ»ÐµÑÐºÐ°Ñ ârecB ÑÐµÐ»Ð¸ÐºÐ°Ð·Ð½Ð°Ñ Ð°ÐºÑÐ¸Ð²Ð½Ð¾ÑÑÑ RecBCD Ð·Ð°Ð¼ÐµÑÐ°ÐµÑÑÑ ÑÐµÐ»Ð¸ÐºÐ°Ð·Ð¾Ð¹ RecQ Ð² Ð¿ÑÐ¾ÑÐµÑÑÐµ Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð¾Ð¹ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ð¸. Ð¢Ð°ÐºÐ¶Ðµ Ð±ÑÐ»Ð¸ Ð¿Ð¾Ð»ÑÑÐµÐ½Ñ ÑÐµÐ·ÑÐ»ÑÑÐ°ÑÑ, ÑÐ²Ð¸Ð´ÐµÑÐµÐ»ÑÑÑÐ²ÑÑÑÐ¸Ðµ Ð¾ ÑÐ½Ð¸Ð¶ÐµÐ½Ð¸Ð¸Â ÑÑÑÐµÐºÑÐ¸Ð²Ð½Ð¾ÑÑÐ¸ Ð¿ÑÐ°Ð¹Ð¼Ð¸ÑÐ¾Ð²Ð°Ð½Ð½Ð¾Ð¹ Ð°Ð´Ð°Ð¿ÑÐ°ÑÐ¸Ð¸ Ð½Ð° ÑÐ¾Ð½Ðµ Ð¸Ð½Ð°ÐºÑÐ¸Ð²Ð°ÑÐ¸Ð¸ ÑÐµÐ»Ð¸ÐºÐ°Ð·Ð½Ð¾Ð¹ Ð°ÐºÑÐ¸Ð²Ð½Ð¾ÑÑÐ¸ ÑÑÐ±ÑÐµÐ´Ð¸Ð½Ð¸ÑÑ RecD ÐºÐ¾Ð¼Ð¿Ð»ÐµÐºÑÐ° RecBCD Ð°Ð¼Ð¸Ð½Ð¾ÐºÐ¸ÑÐ»Ð¾ÑÐ½Ð¾Ð¹ Ð·Ð°Ð¼ÐµÐ½Ð¾Ð¹ K177Q.
description: The tasks of the study: - to obtain strains with deletions âxseA, ârecJ âxseA, ârecQ, ârecB ârecQ; - to test the ability of the ExoVII nuclease to substitute the RecJ exonuclease activity in the process of primed adaptation in cells with the recJ deletion; - to study the ability of the RecQ helicase to substitute the RecBCD helicase activity in the process of primed adaptation in cells with the recB deletion; - to compare the efficiency of adaptation in cells with a point mutation or the recD deletion inactivating the RecD helicase activity. The object of the study is the Escherichia coli KD403 strain with a self-targeting type I-E CRISPR-Cas system, which includes cas genes under the control of inducible promoters and a CRISPR array with a single spacer targeting a protospacer in the yihN gene in the cell genome. The subject of the study is primed adaptation in the type I-E system and the contribution of various repair and recombination genes to the efficiency of this process. The work was carried out in the analytical center of nano- and biotechnologies of Peter the Great St. Petersburg Polytechnic University. We showed that ExoVII substitutes the nuclease activity of RecJ in cells during primed adaptation. RecQ is involved in primed adaptation in ârecB cells. We also demonstrated that the efficiency of primed adaptation is decreased upon inactivation of the RecD helicase activity by the K177Q amino acid substitution.