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Effects of CRISPR/Cas9 dosage on TICAM1 and RBL gene mutation rate, embryonic development, hatchability and fry survival in channel catfish.

Abstract：

:The current study was conducted to assess the effects of microinjection of different dosages of guide RNA (gRNA)/Cas9 protein on the mutation rate, embryo survival, embryonic development, hatchability and early fry survival in channel catfish, Ictalurus punctatus. Guide RNAs targeting two of the channel catfish immune-related genes, toll/interleukin 1 receptor domain-containing adapter molecule (TICAM 1) and rhamnose binding lectin (RBL) genes, were designed and prepared. Three dosages of gRNA/Cas9 protein (low, 2.5 ng gRNA/7.5 ng Cas9, medium, 5 ng gRNA/15 ng Cas9 and high, 7.5 ng gRNA/22.5 ng Cas9) were microinjected into the yolk of one-cell embryos. Mutation rate increased with higher dosages (p < 0.05). Higher dosages increased the mutation frequency in individual embryos where biallelic mutations were detected. For both genes, microinjection procedures increased the embryo mortality (p < 0.05). Increasing the dosage of gRNA/Cas9 protein increased the embryo mortality and reduced the hatching percent (p < 0.05). Embryonic development was delayed when gRNAs targeting RBL gene were injected. Means of fry survival time were similar for different dosages (p > 0.05). The current results lay the foundations for designing gene editing experiments in channel catfish and can be used as a guide for other fish species.

journal_name

Sci Rep

journal_title

Scientific reports

authors

Elaswad A,Khalil K,Ye Z,Liu Z,Liu S,Peatman E,Odin R,Vo K,Drescher D,Gosh K,Qin G,Bugg W,Backenstose N,Dunham R

doi

10.1038/s41598-018-34738-4

subject

Has Abstract

pub_date

2018-11-07 00:00:00

pages

16499

issue

1

issn

2045-2322

pii

10.1038/s41598-018-34738-4

journal_volume

8

pub_type

杂志文章

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