苹果果实大小相关的ARF-Aux/IAA互作组合筛选  

Screening of ARF-Aux/IAA Interaction Combinations Involved in Apple Fruit Size

作　　者：周喆 卞书迅 张恒涛[1] 张瑞萍[1] 高启明[1] 刘珍珍[1] 阎振立[1] ZHOU Zhe;BIAN ShuXun;ZHANG HengTao;ZHANG RuiPing;GAO QiMing;LIU ZhenZhen;YAN ZhenLi(Zhengzhou Fruit Research Institute,Chinese Academy of Agricultural Sciences,Zhengzhou 459000)

机构地区：[1]中国农业科学院郑州果树研究所,郑州459000

出　　处：《中国农业科学》2021年第14期3088-3096,共9页Scientia Agricultura Sinica

基　　金：国家重点研发计划(2018YFD1000106)。

摘　　要：【目的】通过转录组学和生物信息学,在苹果全基因组中对可能互作的MdARF和MdIAA进行鉴定,为明确相关基因功能和解析生长素调控苹果果实大小的分子机理奠定基础。【方法】对野生大果型‘皇家嘎啦’和miRNA172p过表达的转基因小果型‘皇家嘎啦’进行不同发育时期和不同组织材料的转录组测序,对测序结果进行基因的功能注释及差异表达分析。利用转基因小果和野生型大果的转录组数据,筛选在果实发育中表达的苹果MdARFs和MdIAAs基因家族成员,通过逐一在两个家族间计算基因时空表达的相关系数,筛选可能互作的MdARF-MdIAA组合,将从拟南芥基因组中下载的23个ARFs和34个Aux/IAAs、番茄基因组中下载的21个ARFs和25个Aux/IAAs,分别与互作候选MdARFs和MdIAAs进行比对,并进一步构建系统发育树。使用MEME和TBtools对苹果候选互作对中的MdARFs和MdIAAs蛋白进行Motif分析。利用STRING蛋白互作预测数据库进行同源映射,构建苹果中的蛋白-蛋白互作网络,进一步的确认候选互作对,最终得到苹果中通过互作参与果实发育可能性最高的MdARF-MdIAA组合。【结果】分别对野生型‘皇家嘎啦’和miR172OX转基因‘皇家嘎啦’盛花期后两周的全果和盛花期后4周的果皮、果肉和果核进行转录组测序,共生成178.19 Gb的数据量,各项指标均表明,3个生物学重复在所有组织类型上均具有高度一致性。在转录组数据中,共鉴定到38个MdARFs和27个MdIAAs在至少一个文库中的FPKM值大于2,在苹果果实发育时期表达。通过计算Pearson相关系数对表达的MdARFs和MdIAAs两两进行相关性分析,其中8对MdARF-MdIAA的相关系数大于0.9或小于-0.9,作为初步筛选的候选互作组合。将8对组合中的MdARFs和MdIAAs分别与拟南芥和番茄中的ARFs和IAAs进行序列比对并构建系统进化树后发现,MdARF6和MdARF19与起转录激活作用的AtARFs同属一个分支。而MdARF【Objective】The transcriptomics and bioinformatics methods were used to carry out the genome-wide potential interacting MdARFs and MdIAAs pairs screening,so as to build a foundation for clarifying the function of related genes and elucidating the molecular mechanism underlying auxin-regulated apple fruit size.【Method】Two apple genotypes,Royal Gala(WT)and 35S::miRNA172p transgenic Royal Gala(miR172OX)were used as test materials in this study.Fruit materials from different developmental stages and tissue types were collected for both genotypes and were subjected to transcriptome sequencing.Clean reads were aligned to the reference genome and the differential expression analysis was performed.Based on the transcriptome data obtained by sequencing the transgenic small fruit and wild-type large fruit,the pairwise expression analysis was performed across MdARFs and MdIAAs families.The amino acid sequences of 23 ARFs and 34 Aux/IAAs were downloaded from Arabidopsis genome,and 21 ARFs and 25 Aux/IAAs were downloaded from tomato genome,which were further compared with candidate MdARFs and MdIAAs to construction phylogenetic trees.The MEME and TBtools were used to carry out the Motif analysis for candidate MdARFs and MdIAAs.Pairs with high interacting possibilities were further confirmed by a protein-protein interacting network constructed in apple to finalize combinations with the highest probability of involvement in fruit development.【Result】The whole fruit at 2 weeks post full bloom and the fruit skin,fruit flesh and fruit core at 4 WPFB were collected from WT and miR172OX,respectively.To achieve research objectives,transcriptome sequencing was carried out.A total of 178.19 Gb paired-end reads of 125 bp/150 bp were generated.All indexes indicated that the three biological replicates had highly consistent transcriptome profiles across all tissue types.FPKM values in at least one library was over 2 were used as a standard to eliminate the low expressed genes,so a total of 38 MdARFs and 27 MdIAAs were expressed

关 键 词：苹果 ARF AUX/IAA 互作筛选 

分 类 号：S661.1[农业科学—果树学]