Target-specific mutations efficiency at multiple loci of CRISPR/Cas9 system using one sgRNA in soybean
Soybean has a palaeopolyploid genome with nearly 75% of the genes present in multiple copies. Although the CRISPR/Cas9 system has been employed in soybean to generate site-directed mutagenesis, a systematical assessment of mutation efficiency of the CRISPR/Cas9 system for the multiple-copy genes is...
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Veröffentlicht in: | Transgenic research 2021-02, Vol.30 (1), p.51-62 |
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Sprache: | eng |
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Zusammenfassung: | Soybean has a palaeopolyploid genome with nearly 75% of the genes present in multiple copies. Although the CRISPR/Cas9 system has been employed in soybean to generate site-directed mutagenesis, a systematical assessment of mutation efficiency of the CRISPR/Cas9 system for the multiple-copy genes is still urgently needed. Here, we successfully optimize one sgRNA CRISPR/Cas9 system in soybean by testing the efficiency, pattern, specificity of the mutations at multiple loci of
GmFAD2
and
GmALS
. The results showed that simultaneous site-directed mutagenesis of two homoeologous loci by one sgRNA, the mutation frequency in the T0 generation were 64.71% for
GmPDS
, 60.0% for
GmFAD2
and 42.86% for
GmALS
, respectively. The chimeric and heterozygous mutations were dominant types. Moreover, association of phenotypes with mutation pattern at target loci of
GmPDS11
and
GmPDS18
could help us further demonstrate that the CRISPR/Cas9 system can efficiently generate target specific mutations at multiple loci using one sgRNA in soybean, albeit with a relatively low transformation efficiency. |
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ISSN: | 0962-8819 1573-9368 |
DOI: | 10.1007/s11248-020-00228-5 |