Time origin and structural analysis of the induced CRISPR/cas9 megabase-sized deletions and duplications involving the Cntn6 gene in mice

Inna E. Pristyazhnyuk, Julia Minina, Alexey Korablev, Irina Serova, Veniamin Fishman, Maria Gridina, Timofey S. Rozhdestvensky, Leonid Gubar, Boris V. Skryabin, Oleg L. Serov

Результат исследования: Научные публикации в периодических изданияхстатья

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Аннотация

In a previous study using one-step CRISPR/Cas9 genome editing in mouse zygotes, we created five founders carrying a 1,137 kb deletion and two founders carrying the same deletion, plus a 2,274 kb duplication involving the Cntn6 gene (encoding contactin-6). Using these mice, the present study had the following aims: (i) to establish stage of origin of these rearrangements; (ii) to determine the fate of the deleted DNA fragments; and (iii) to estimate the scale of unpredicted DNA changes accompanying the rearrangements. The present study demonstrated that all targeted deletions and duplications occurred at the one-cell stage and more often in one pronucleus only. FISH analysis revealed that there were no traces of the deleted DNA fragments either within chromosome 6 or on other chromosomes. These data were consistent with the Southern blot analysis showing that chromosomes with deletion often had close to expected sizes of removed DNA fragments. High-throughput DNA sequencing of two homozygotes for duplication demonstrated that there were no unexpected significant or scale DNA changes either at the gRNA and joint sites or other genome sites. Thus, our data suggested that CRISPR/Cas9 technology could generate megabase-sized deletions and duplications in mouse gametes at a reasonably specific level.

Язык оригиналаанглийский
Номер статьи14161
Число страниц1
ЖурналScientific Reports
Том9
Номер выпуска1
DOI
СостояниеОпубликовано - 2 окт 2019

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