Map-Cloning Of OsDLT10,A Low Tiller And Dwarf Gene In Rice

The tillering number,which formation and growth process are regulated by gnentic and external factors such as hormones,temperature,moisture,and light,etc,affect rice yield through directly impacting the number of effective panicles.The researches of genes related to the tillering number are conducive to elucidate the molecular mechanism of the axillary buds formation and development,which is of great importance for the construction of ideal plant types in rice.In this study,a low tillering and dwarf mutant osdlt10(dwarf and low tillering 10)was obtained from Zhonghui8015(Wild Type,WT)by Ethyl Methanesulfonate(EMS)mutagenesis.The effects of OsDLT10 on axillary bud formation,development and related regulatory pathways were analysised in the hope of providing a potencial gene resource and theoretical basis for the breeding application of plant types in rice.The main results are as follows:1.The phenotype of osdlt10 mutant lasted throughout the whole growth period.The growth retardation and shallow root first appeared at the seedling stage.In addition,the mutant exhibited low tillering,dwarf and leaf tip curling at early tillering stage.The dwarf and low tillering phenotype gradually became more significant with the growth and development.At mature stage,besides the low tillering number and decreased plant height,the grain width,seed setting rate and number of branches also decreased significantly in the mutant.Cytological observation results of axillary bud indicated that the formation of axillary bud was defective and grew slowly.Furthermore,OsDLT10 also affects cell size in different tissues such as axillary bud,stem,leaf,glume,root tip.2.Genetic analysis showed that the osdlt10 phenotype is controlled by a single recessive nuclear gene.The osdlt10 was finely located on the long arm of chromosome 10 flanked by wxx-17 and wxx-53,with a physical distance of about 57.78 kb harboring 10 Open Reading Frames(ORFs)through the map-based cloning method.Sequencing results revealed that a single base(G)deletion within the 9th ORF,resulted in a premature translation termination after a frameshift mutation in the encoded amino acid.Functional complementation and CRISPR/Cas9 knockout demonstrasted that the DUF630/632 gene LOC_Os10g41310 is the responsible gene for the osdlt10 phenotype.Subcellular localization displayed OsDLT10 was localized a dotted-like organelle in the cytoplasm.3.The expression pattern analysis results suggested that OsDLT10 was highly expressed in stem nodes,axillary bud bases,leaf sheath bases and pulvinus.In addition,the expression level of OsDLT10 showed a gradual upward trend throughout the growth and development process.Hormone treatment results revealed that high exogenous auxin concentration could inhibit the expression of OsDLT10,and the endogenous auxin content of the stem nodes bases and axillary bud bases of osdlt10 mutants was significantly reduced.The above results indicated that the expression of OsDLT10 may be related to auxin.The qPCR analysis of auxin-related genes suggested that the expression levels of auxin transport genes(PINs)and auxin early response genes(IAAs)in the osdlt10 mutant were significant up-regulated,indicating that the recessive mutation of this gene may influence auxin polarity transport and local auxin content.4.qPCR analysis of tillering number-related genes revealed that the expression of WUS genes(MOC3,DWT1,WOX4)and FON1 were significantly down-regulated,while the expression levels of MOC1 and OsTB1 were significantly up-regulated,suggesting that the formation and growth of axillary buds may be influenced by WUS-CLV factors.As a result,the tillering of the osdlt10 mutant decreased significantly.