TY - JOUR

T1 - On-target CRISPR/CAS9 activity can cause undesigned large deletion in mouse zygotes

AU - Korablev, Alexey

AU - Lukyanchikova, Varvara

AU - Serova, Irina

AU - Battulin, Nariman

N1 - Publisher Copyright: © 2020 by the authors. Licensee MDPI, Basel, Switzerland.

PY - 2020/5/20

Y1 - 2020/5/20

N2 - Genome engineering has been tremendously affected by the appearance of the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9)-based approach. Initially discovered as an adaptive immune system for prokaryotes, the method has rapidly evolved over the last decade, overtaking multiple technical challenges and scientific tasks and becoming one of the most effective, reliable, and easy-to-use technologies for precise genomic manipulations. Despite its undoubtable advantages, CRISPR/Cas9 technology cannot ensure absolute accuracy and predictability of genomic editing results. One of the major concerns, especially for clinical applications, is mutations resulting from error-prone repairs of CRISPR/Cas9-induced double-strand DNA breaks. In some cases, such error-prone repairs can cause unpredicted and unplanned large genomic modifications within the CRISPR/Cas9 on-target site. Here we describe the largest, to the best of our knowledge, undesigned on-target deletion with a size of ~293 kb that occurred after the cytoplasmic injection of CRISPR/Cas9 system components into mouse zygotes and speculate about its origin. We suppose that deletion occurred as a result of the truncation of one of the ends of a double-strand break during the repair.

AB - Genome engineering has been tremendously affected by the appearance of the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9)-based approach. Initially discovered as an adaptive immune system for prokaryotes, the method has rapidly evolved over the last decade, overtaking multiple technical challenges and scientific tasks and becoming one of the most effective, reliable, and easy-to-use technologies for precise genomic manipulations. Despite its undoubtable advantages, CRISPR/Cas9 technology cannot ensure absolute accuracy and predictability of genomic editing results. One of the major concerns, especially for clinical applications, is mutations resulting from error-prone repairs of CRISPR/Cas9-induced double-strand DNA breaks. In some cases, such error-prone repairs can cause unpredicted and unplanned large genomic modifications within the CRISPR/Cas9 on-target site. Here we describe the largest, to the best of our knowledge, undesigned on-target deletion with a size of ~293 kb that occurred after the cytoplasmic injection of CRISPR/Cas9 system components into mouse zygotes and speculate about its origin. We suppose that deletion occurred as a result of the truncation of one of the ends of a double-strand break during the repair.

KW - CRISPR/Cas9

KW - Cytoplasmic microinjections

KW - Kit knockout mice

KW - Large deletion

KW - On-target deletions

KW - Truncation

KW - Zygotic microinjections

KW - CRISPR

KW - VARIANTS

KW - KIT GENE

KW - DOMINANT-WHITE

KW - large deletion

KW - on-target deletions

KW - GENOME

KW - cytoplasmic microinjections

KW - ONE-STEP GENERATION

KW - DELIVERY

KW - zygotic microinjections

KW - CRISPR-CAS9 NUCLEASES

KW - CAS9 RIBONUCLEOPROTEIN

KW - MICE

KW - Cas9

KW - MUTATIONS

KW - truncation

UR - http://www.scopus.com/inward/record.url?scp=85085265191&partnerID=8YFLogxK

U2 - 10.3390/ijms21103604

DO - 10.3390/ijms21103604

M3 - Article

C2 - 32443745

AN - SCOPUS:85085265191

VL - 21

JO - International Journal of Molecular Sciences

JF - International Journal of Molecular Sciences

SN - 1661-6596

IS - 10

M1 - 3604

ER -