Generation of a new thermo-sensitive genic male sterile rice line by targeted mutagenesis of TMS5 gene through CRISPR/Cas9 system

Abstract

BackgroundTwo-line hybrid rice with high yield potential is increasingly popular and the photo- and temperature-sensitive male sterile line is one of the basic components for two-line hybrid rice breeding. The development of male sterile lines through conventional breeding is a lengthy and laborious process, whereas developing thermo-sensitive genic male sterile (TGMS) lines for two-line hybrid breeding by editing a temperature-sensitivity gene by CRISPR/Cas9 is efficient and convenient.ResultsHere, thermo-sensitive genic male sterility (TGMS) was induced by employing the CRISPR/Cas9 gene editing technology to modify the gene TMS5. Two TGMS mutants, tms5–1 and tms5–2, both lacking any residual T-DNA, were generated in the indica rice cultivar Zhongjiazao17 (cv. YK17) background. When grown at a sub-optimal temperature (22 °C), both mutants produced viable pollen and successfully produced grain through self-fertilization, but at temperatures 24 and 26 °C, their pollen was sterile and no grain was set. F1 hybrids derived from the crosses between YK17S (tms5–1) and three different restorer lines outperformed both parental lines with respect to grain yield and related traits.ConclusionThe YK17S generated by CRISPR/Cas9 system was proved to be a new TGMS line with superior yield potential and can be widely utilized in two-line hybrid breeding of indica rice.Electronic supplementary materialThe online version of this article (10.1186/s12870-019-1715-0) contains supplementary material, which is available to authorized users.

Design of high-oleic tobacco (Nicotiana tabacum L.) seed oil by CRISPR-Cas9-mediated knockout of NtFAD2–2

“…T0 generation lines were selected on media with 50 mg/L kanamycin. Mutation selection was performed as previous reported with some modification [79]. Briefly, genomic DNA was extracted from the leaves of the kanamycinresistant T0 transgenic plants using the Plant DNA Isolation Kit (Foregene, China).…”

Base Editing: The Ever Expanding Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) Tool Kit for Precise Genome Editing in Plants

“…The development of genome-editing technology has revolutionized agricultural studies, enabling the improvement of different plant traits [1][2][3][4][5]. Various tools have been widely used for precise genome editing in plants, including clustered regularly interspaced short palindromic repeats (CRISPR) based gene editing techniques, such as CRISPR/CRISPR associated protein 9 (Cas9), CRISPR/Cas12a (Cpf1), and CRISPR/Cas13 [6][7][8][9], owing to their reliability.…”

“…Various tools have been widely used for precise genome editing in plants, including clustered regularly interspaced short palindromic repeats (CRISPR) based gene editing techniques, such as CRISPR/CRISPR associated protein 9 (Cas9), CRISPR/Cas12a (Cpf1), and CRISPR/Cas13 [6][7][8][9], owing to their reliability. All these techniques have certain advantages and disadvantages and have become more precise, efficient, and robust in recent years [1,[10][11][12][13]. The formation of DNA double-strand breaks (DSBs) at target loci is one of the primary features of CRISPR/Cas genome editing.…”

Targeted genome editing inNicotiana tabacumusing inducible CRISPR/Cas9 system

“…Among them, the CRISPR/Cas9 system is predominantly used for TGE due to its simplicity, high efficiency and versatility [8]. In most cases, the Cas9 encoding gene is driven by constitutive promoters, such as CaMV 35S and ubiquitin promoters [9,10,11,12,13]. However, constitutive expression of Cas9 might be harmful to cells, or increase the risk of off-target effect [14,15].…”