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Conversion of a normal maize hybrid into a waxy version using in vivo CRISPR/Cas9 targeted mutation activity

查看全文 作　　者：Xiantao [1,2]Qi;Hao [1,2]Wu;Haiyang [1]Jiang;Jinjie [2]Zhu;Changling [2]Huang;Xin [1]Zhang;Changlin [2]Liu;Beijiu [1]Cheng 高影响力作者 机构地区：[1]Anhui Agricultural University,Hefei 230036,Anhui,China;[2]Institute of Crop Sciences,Chinese Academy of Agricultural Sciences/National Key Facility for Crop Gene Resources and Genetic Improvement,Beijing 100081,China高影响力机构 出　　处：《The Crop Journal》索引2020年第8卷第3期,共9页高影响力期刊 基　　金：supported the National Transgenic Science and Technology Program(2019ZX08010-003);the National Natural Science Foundation of China(31771808);the National Key Research and Development Program of China(2016YFD0101803);the Key Area Research and Development Program of Guangdong Province(2018B020202008);Beijing Municipal Science and Technology Commission(D171100007717001);National Engineering Laboratory for Crop Molecular Breeding。 摘　　要：Waxy maize is a specialty maize that produces mainly amylopectin starch with special food or industrial values. The objective of this study was to overcome the limitations of wx mutant allele acquisition and breeding efficiency by conversion of parental lines from normal to waxy maize. The intended mutation activity was achieved by in vivo CRISPR/Cas9 machinery involving desired-target mutation of the Wx locus in the ZC01 background,abbreviated as ZC01-DTM^(wx). Triple selection was applied to segregants to obtain high genome background recovery with transgene-free wx mutations. The targeted mutation was identified, yielding six types of mutations among progeny crossed with ZC01-DTM^(wx).The amylopectin contents of the endosperm starch in mutant lines and hybrids averaged94.9%, while those of the wild-type controls were significantly(P < 0.01) lower, with an average of 76.9%. Double selection in transgene-free lines was applied using the Bar strip test and Cas9 PCR screening. The genome background recovery ratios of the lines were determined using genome-wide SNP data. That of lines used as male parents was as high as98.19% and that of lines used as female parents was as high as 86.78%. Conversion hybrids and both parental lines showed agronomic performance similar to that of their wild-type counterparts. This study provides a practical example of the efficient extension of CRISPR/Cas9 targeted mutation to industrial hybrids for transformation of a recalcitrant species. 关 键 词：Conversion of a normal maize hybrid into a waxy version using in vivo CRISPR/Cas9 targeted mutation activity CRISPR
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