Re-domestication of potato into an inbred line-based diploid crop propagated by seed represents a promising alternative to traditional clonal propagation of tetraploid potato, but self-incompatibility has hindered the development of inbred lines. To address this problem, we created self-compatible diploid potatoes by knocking out the self-incompatibility gene S-RNase using the CRISPR-Cas9 system. This strategy opens new avenues for diploid potato breeding and will also be useful for studying other self-incompatible crops.
Clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins (CRISPR/Cas) is an acquired immune system found in bacteria and archaea that can specifically silence or degrade a foreign single or double strand nucleic acid to protect it from infection. In recent years, the CRISPR/Cas9 system has rapidly been evolved into a genome editing technology, in which the Cas9 endonuclease can be targeted to specific DNA sequences by guide RNAs (gRNAs) that are easily programmable. Due to simplicity, specificity and high efficiency, CRISPR/Cas9 is gradually replacing other gene editing technologies and has been implemented in basic and applied plant sciences to boost yield, regulate metabolic process, and increase stress resistance in different varieties. In current review, we introduced its application scope in scientific research and practical application. We summarized the procedure of target plant generation by CRISPR/Cas9 method. We mainly reviewed the applications of CRISPR/Cas9 and its recent advances in model plants and other crop plants, attempting to provide a related general information to researchers. Further, we also included the inadequacies and concerns of CRISPR/Cas9 that have emerged so far.
Reactive oxygen species (ROS), especially hydrogen peroxide, H2O2, act as signaling molecules to widely mediate growth, development, and stress response of plants. In the present study, internal ROS accumulation, effects of exogenous H2O2 treatment, the expression of the key tuberization-related genes, and the effect of knockout of Solanum tuberosum self-pruning 6A (StSP6A) on H2O2-induced tuber formation were investigated to elucidate whether and how H2O2 is involved in induction and formation of potato tubers using two diploid landraces, Solanum phureja and S. ajanhuiri. The results showed that there was a significant accumulation of ROS (including H2O2, superoxide anion, O2−, and total ROS) during tuber induction and formation in stolons/tubers, especially in the hook-like subapical part of stolons prior to tuberization, as detected by staining observation and quantitative measurement. Furthermore, exogenous H2O2 treatment significantly enhanced percentage of tuber formation. By contrast, addition of either the ROS inhibitor diphenyleneiodonium chloride (DPI) or H2O2 scavenger catalase (CAT) resulted in a decline of tuber formation. In addition, expression analysis of nine key tuberization-related genes demonstrated that the H2O2-induced tuberization could be associated with H2O2-controlled regulation of these tuberization- and signaling-pathway-related genes, especially StSP6A, which was dramatically up-regulated during the early stage of tuber induction and H2O2 treatment. When StSP6A was knocked out by CRISPR-Cas9-mediated genome editing, the tuberization frequency of StSP6A null-mutants became significantly lower at various H2O2 concentration treatments. These findings indicate that H2O2 accumulation in stolons might play an important role by acting as a signaling molecule to initiate tuber induction, H2O2-induced tuber formation is triggered by regulating the tuberization-related gene expression and activating signal transduction pathways, and StSP6A is a pivotal player in H2O2-induced tuber formation in potato.
Reactive oxygen species (ROS, especially hydrogen peroxide, H2O2) act as signaling molecules to mediate growth, development and stress response of plants. However, whether and how H2O2 is involved in induction and formation of potato tubers is little known. The present study showed that there was an obvious accumulation of ROS (including H2O2, O2·- and total ROS) during tuber induction and formation in stolons/tubers, especially in the hook-like subapical part of stolons prior to tuberization detected by staining observation and quantitative measurement. Furthermore, exogenous H2O2 treatment significantly enhanced percentage of tuber formation. By contrast, addition of either the ROS inhibitor diphenyleneiodonium chloride (DPI) or H2O2 scavenger catalase (CAT) resulted in a decline of tuber formation. In addition, expression analysis of 9 key tuberization-related genes demonstrated that the H2O2-induced tuberization could be associated with H2O2-controlled regulation of these tuberization- and signaling pathway-related genes, especially SELF-PRUNING 6A gene (StSP6A), which was dramatically up-regulated during early stage of tuber induction and H2O2 treatment. When StSP6A was knocked out by CRISPR-Cas9-mediated genome editing, the tuberization frequency of StSP6A null-mutants became extremely lower under H2O2 treatments. These findings indicate that H2O2 accumulation in stolons might plays an important role acting as a signaling molecule to initiate tuber induction, H2O2-induced tuber formation is triggered by regulating the tuberization-related gene expression and activating signal transduction pathways, and StSP6A is a pivotal player in the H2O2-induced tuber formation in potato.
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