Effects Of Auxin Repressed Protein Gene OsARP1 In Rice Germination And Seedling Growth

Auxin repressed protein genes,ARPs,downregulated by IAA signal,play important roles in plant growth and development,resistance to diseases and stresses,and seed dormancy.However,no studied was reported in rice Auxin repressed protein gene,OsARP1.In this work,we generated overexpression lines and exon-specific knockout mutants of OsARP1 to study the difference in seed germination and seedling growth of these transgenic plants.The main results are as follows:1.191 OsARP1 transgenic regenerated plants were obtained,of which,56 were overexpression plants with a positive rate of 93.33%.27 of the overexpression rice plant showed over doubled OsARP1expression levels over double of the expression levels.2.The poisitve rate of OsARP1 promoter-sepcific knockout plants was 93.62%（44 promoter-sepcific knockout plants in 47 plants）.Mutants in 28 plants were identified in target site 1,and 13 of which were homozygous.27 in target site 2 with 8homozygous plants.Double mutants were identified in 24 plants with 8 homozygous plants.In total of 85 mutations were identified,of which,30 were base substitutions with 26 single base substitutions.36 were deletions（14 single base deletions,11multiple base deletions）.19 were base insertions（14 single base insertions,2 multiple base insertions）.We identified a 27.06%homozygoue rate.Variation in the OsARP1promoter significantly reduced OsARP1 expression level manifested by qRT-PCR analysis.3.The poisitve rate of OsARP1 exon-sepcific knockout plants was 94.00%（47exon-sepcific knockout plants in 50 plants）.Mutants in 28 plants were identified in target site 1,and 10 of which were homozygous.30 in target site 2 with 8homozygous plants.Double mutants were identified in 28 plants with 6 homozygous plants.In total of 71 mutations were identified,of which,15 were base substitutions with 8 single base substitutions.41were deletions（3single base deletions,33 multiple base deletions）.15 were base insertions（7 single base insertions,2 multiple base insertions）.We identified a 22.53%homozygoue rate.4.3 homogeneous mutants were selected as parents from T₀ promoter-and exon-specific knockout lines distinctly for T₁ plants.Edited target sites were successfully identified in 31 promoter specific knockouts and 35 exon specific knockouts,29 and34 of which were identical to their T₀ parents,with only 3 brand new mutations.95.52%of the mutations in T₁ remained identical to those in T₀.5.The rate of seed germination and seedling height were significantly lower in promoter-and exon-specific knockout plants compared to WTs,suggesting that rice growth and development could be impacted by downregulation or loss of function of OsARP1.Our work provide good material for further study on the function of OsARP genes in rice growth and development,resistance to diseases and stresses.