Gibberellin （GA） 2-oxidase plays a key role in the GA catabolic pathway through 2β-hydroxylation.In the present study,we isolated a CaMV 35S-enhancer activation tagged mutant,H032.This mutant exhibited a dominant d...Gibberellin （GA） 2-oxidase plays a key role in the GA catabolic pathway through 2β-hydroxylation.In the present study,we isolated a CaMV 35S-enhancer activation tagged mutant,H032.This mutant exhibited a dominant dwarf and GA-deficient phenotype,with a final stature that was less than half of its wild-type counterpart.The endogenous bioactive GAs are markedly decreased in the H032 mutant,and application of bioactive GAs （GA3 or GA4） can reverse the dwarf phenotype.The integrated T-DNA was detected 12.8 kb upstream of the OsGA2ox6 in the H032 genome by TAIL-PCR.An increased level of OsGA2ox6 mRNA was detected at a high level in the H032 mutant,which might be due to the enhancer role of the CaMV 35S promoter.RNAi and ectopic expression analysis of OsGA2ox6 indicated that the dwarf trait and the decreased levels of bioactive GAs in the H032 mutant were a result of the up-regulation of the OsGA2ox6 gene.BLASTP analysis revealed that OsGA2ox6 belongs to the class III of GA 2-oxidases,which is a novel type of GA2ox that uses C20-GAs （GA12 and/or GA53） as the substrates.Interestingly,we found that a GA biosynthesis inhibitor,paclobutrazol,positively regulated the OsGA2ox6 gene.Unlike the over-expression of OsGA2ox1,which led to a high rate of seed abortion,the H032 mutant retained normal flowering and seed production.These results indicate that OsGA2ox6 mainly affects plant stature,and the dominant dwarf trait of the H032 mutant can be used as an efficient dwarf resource in rice breeding.展开更多
The CRISPR/Cas(clustered regularly in terspaced short palindromic repeats/CRISPR-associated protein)system has been widely used in genome editing,epigenetic modification,and other applications,because it is a precise,...The CRISPR/Cas(clustered regularly in terspaced short palindromic repeats/CRISPR-associated protein)system has been widely used in genome editing,epigenetic modification,and other applications,because it is a precise,inexpensive,powerful and easy-to?use editing tool(Zhang et al.,2019).However,the scope of genome editing is limited by a short protospacer adjacent motif(PAM)specific sequence flanking the target site.Streptococcus pyogenes Cas9(5^Cas9),the most widely used Cas,strongly recognizes NGG PAM and partially recognizes NAG PAM in rice and is restricted to target the sites with these motifs(Anders et al.,2014;Meng et al.,2018).展开更多
MSH5, a meiosis-specific member of the MutS-homolog family, is required for normal level of recombination in budding yeast, mice, Caenorhabditis elegans, and Arabidopsis. Here, we report the identification and charact...MSH5, a meiosis-specific member of the MutS-homolog family, is required for normal level of recombination in budding yeast, mice, Caenorhabditis elegans, and Arabidopsis. Here, we report the identification and characterization of its rice homolog, OsMSH5, and demonstrate its function in rice meiosis. Five independent Osmsh5 mutants exhibited normal vegetative growth and severe sterility. The synaptonemal complex is well installed in Osmsh5, while the chiasma frequency is greatly reduced to approximately 10% of that observed in the wild-type, leading to the homologous non- disjunction and complete sterile phenotype. OsMSH5 is predominantly expressed in panicles. Immunofluorescence studies indicate that OsMSH5 chromosomal localization is limited to the early meiotic prophase I. OsMSH5 can be loaded onto meiotic chromosomes in Oszip4, Osmer3, and hellO. However, those ZMM proteins cannot be localized normally in the absence of OsMSH5. Furthermore, the residual chiasmata were shown to be the least frequent among the zmm mutants, including Osmer3, Oszip4, hellO, and Osmsh5. Taken together, we propose that OsMSH5 functions upstream of OsZIP4, OsMER3, and HEIl0 in class I crossover formation.展开更多
基金supported by grants from the Ministry of Sciences and Technology of China (No. 2005CB120805 and 2006AA10A101)the National Natural Science Foundation of China (No. 30621001 and 30871512)
文摘Gibberellin （GA） 2-oxidase plays a key role in the GA catabolic pathway through 2β-hydroxylation.In the present study,we isolated a CaMV 35S-enhancer activation tagged mutant,H032.This mutant exhibited a dominant dwarf and GA-deficient phenotype,with a final stature that was less than half of its wild-type counterpart.The endogenous bioactive GAs are markedly decreased in the H032 mutant,and application of bioactive GAs （GA3 or GA4） can reverse the dwarf phenotype.The integrated T-DNA was detected 12.8 kb upstream of the OsGA2ox6 in the H032 genome by TAIL-PCR.An increased level of OsGA2ox6 mRNA was detected at a high level in the H032 mutant,which might be due to the enhancer role of the CaMV 35S promoter.RNAi and ectopic expression analysis of OsGA2ox6 indicated that the dwarf trait and the decreased levels of bioactive GAs in the H032 mutant were a result of the up-regulation of the OsGA2ox6 gene.BLASTP analysis revealed that OsGA2ox6 belongs to the class III of GA 2-oxidases,which is a novel type of GA2ox that uses C20-GAs （GA12 and/or GA53） as the substrates.Interestingly,we found that a GA biosynthesis inhibitor,paclobutrazol,positively regulated the OsGA2ox6 gene.Unlike the over-expression of OsGA2ox1,which led to a high rate of seed abortion,the H032 mutant retained normal flowering and seed production.These results indicate that OsGA2ox6 mainly affects plant stature,and the dominant dwarf trait of the H032 mutant can be used as an efficient dwarf resource in rice breeding.
基金This work was supported by the National Transgenic Science and Technology Program(2019ZX08010-001,2019ZX08010-003 and 2016ZX08001004-002)the Agricultural Science and Technology Innovation Program of Chinese Academy of Agricultural Sciences.
文摘The CRISPR/Cas(clustered regularly in terspaced short palindromic repeats/CRISPR-associated protein)system has been widely used in genome editing,epigenetic modification,and other applications,because it is a precise,inexpensive,powerful and easy-to?use editing tool(Zhang et al.,2019).However,the scope of genome editing is limited by a short protospacer adjacent motif(PAM)specific sequence flanking the target site.Streptococcus pyogenes Cas9(5^Cas9),the most widely used Cas,strongly recognizes NGG PAM and partially recognizes NAG PAM in rice and is restricted to target the sites with these motifs(Anders et al.,2014;Meng et al.,2018).
基金This work was supported by grants from the Ministry of Sciences and Technology of China （2011CB944602 and 2012AA10A301）the State Key Laboratory of Plant Genomics of China （2012A0527）and the National Natural Science Foundation of China （31160223 and 31230038）. No conflict of interest declared.
文摘MSH5, a meiosis-specific member of the MutS-homolog family, is required for normal level of recombination in budding yeast, mice, Caenorhabditis elegans, and Arabidopsis. Here, we report the identification and characterization of its rice homolog, OsMSH5, and demonstrate its function in rice meiosis. Five independent Osmsh5 mutants exhibited normal vegetative growth and severe sterility. The synaptonemal complex is well installed in Osmsh5, while the chiasma frequency is greatly reduced to approximately 10% of that observed in the wild-type, leading to the homologous non- disjunction and complete sterile phenotype. OsMSH5 is predominantly expressed in panicles. Immunofluorescence studies indicate that OsMSH5 chromosomal localization is limited to the early meiotic prophase I. OsMSH5 can be loaded onto meiotic chromosomes in Oszip4, Osmer3, and hellO. However, those ZMM proteins cannot be localized normally in the absence of OsMSH5. Furthermore, the residual chiasmata were shown to be the least frequent among the zmm mutants, including Osmer3, Oszip4, hellO, and Osmsh5. Taken together, we propose that OsMSH5 functions upstream of OsZIP4, OsMER3, and HEIl0 in class I crossover formation.
