- transcriptome of olecranon-type traits in peach ( Prunus persica L. - Background: The top of the olecranon honey peach ( Prunus persica L. - To explore the genetic mechanism of olecranon formation, we performed full-length transcriptome sequencing analysis of olecranon and round peaches as well as a genome-wide association study of the association of olecranon-type trait loci.. - The genes were within 1 Mb up- or down-stream of the main genome-wide association study locus for olecranon-type traits.. - It is considered as one of the best peach varieties and is very popular in China because it is sweet, juicy, and has a crisp texture [4].. - The flat shape is determined by a single gene, S/s, in the distal part of. - The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. - If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. - Full list of author information is available at the end of the article. - In addition, differences in the fruit cell number, cell size, and gene expression patterns between the round and flat fruit varieties and the oblate shape of peach fruits are mainly determined by regulating the production of cells in the vertical direction during early fruit development [2]. - The Oxford Nanopore Technologies (ONT, Oxford, UK) sequen- cing method was used to study differences in the transcription of genes affecting the olecranon trait to narrow the range of related genes and provide a basis for further studies of these genes.. - The average number of clean reads of the samples was and the average mapping rate between the samples and reference gene was 97.38 % (Supplemental Table 2). - Averages of 231,407 and 121,336 SNPs were the transition type and transversion type, respectively, and the average SNP ratio of the heterozygous types was 63.39 % (Supplemental Table 3). - 1 Geographic distribution, linkage disequilibrium (LD) decay of 456 peach accessions and genome-wide association scan for olecranon-type traits. - GWAS of olecranon-type traits and prediction of candidate genes. - In this study, we counted 296 types of peaches out of the 456 germplasm resources, among which 155 had olecranon and 141 had no olecranon. - To explore the potential of GWAS to identify causal genes of olecranon-type traits, we used the FaST-LMM model to conduct an associ- ation study of olecranon-type traits. - The locus defined by the SNP at bp was strongly associated with olecranon-type traits (Fig. - We selected 38 genes within 1 Mb up- or down-stream of the main GWAS locus for olecranon-type traits (Supplemental Table 6). - Through Swiss-PROt database annotation, we found that 20 genes are also expressed in the Arabidopsis genome. - This variation was highly associated with an A/T polymorphism detected in the fifth intron of LOC18772384 on NC_034014.1.. - 2B), and was found to have an alteration related to fruit- shape in the bud sport. - Moreover, other branches bore olecranon-type fruits (Fig. - The results showed that the olecranon-type fruit weighed g, and had a longitudinal diameter of cm and transverse diameter of cm. - According to the t-test, the quality indicators of the two types of peaches differed significantly (P <. - Notably, both the weight and vertical and horizontal diameters of olecranon-type fruit were larger than those of round-type fruit. - However, the olecranon- type fruit shape index was smaller than that of round-type fruit. - After removing ribosomal RNA, the numbers of full-length reads in the. - 2 Phenotypic characterization of olecranon honey peach and round peach. - (A) Sampled peach trees, (B) round peach, and (C) olecranon honey peach. - Table 2 Comparison of quality indices of the two peach fruits Project Olecranon-type fruits Round-type fruits. - were 5782 differentially expressed genes (DEGs) in the two groups, including 3293 up-regulated genes and 2489 down-regulated genes with P-values <. - There were 11,239 differentially expressed transcripts (DETs) in the two groups, including 6294 up-regulated transcripts and 4945 down-regulated transcripts with P-values <. - To identify genes important for olecranon-type traits, we annotated the DEGs and DETs using the Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Eukaryotic Orthologous Groups (KOG), Clusters of Orthologous Groups (COG), and eggNOG databases. - In the bio- logical process classification, genes and transcripts. - In the cellular component category, the main subcategories were cell and cell part. - Catalytic activity and binding were the major subcategories in the biological process classification.. - There were 3154 DEGs and 6222 DETs annotated in the KOG database (Supplementary Table 9). - DEGs and DETs were most enriched in the eggNOG database, showing numbers of 5299 and 10,140, respectively (Supplementary Table 9).. - Each point in the differential expression volcano chart represents a gene, and the abscissa represents the logarithm of the multiple of the difference in expression of a certain gene in the two samples. - the ordinate represents the negative logarithm of statistical significance of the change in gene expression. - A larger absolute value of the abscissa indicates a greater fold-difference in expression between the two samples. - a larger ordinate value indicates more significant differential expression and greater reliability of the DEGs obtained by screening. - The green dots in the figure represent down-regulated DEGs, red dots represent up- regulated DEGs, and black dots represent non-DEGs. - Selection of candidate genes for olecranon-type traits First, we selected 38 genes within 1 Mb up- or down- stream of the main GWAS locus for olecranon-type traits (Supplemental Table 6). - The expression of 38 genes in round-type fruits and olecranon-type fruits is shown in Supplemental Tables 10, among which 8 were. - Eight DEGs were selected as candidates affecting olecranon-type traits, including 4 highly expressed genes in olecranon honey peach (LOC18775282, LOC18772209, LOC18773929, LOC18773401) and 4 highly expressed genes in round peach (LOC18773079, LOC18773525,. - The abscissa represents the GO classification, left of the ordinate represents the percentage of the number of genes, and right represents the number of genes. - LOC18773067, LOC18775244) according to the expres- sion pattern shown in the heatmap (Fig. - We selected 80 transcripts with 1 Mb up- or down- stream of the main GWAS locus for olecranon-type traits. - Among the 80 transcripts, 39 were not expressed or showed very low expression in the round-type fruits and olecranon-type fruits and 11 transcripts (FDR <. - According to expression pattern in the heat- map (Fig. - Thus, we selected 10 genes and one new gene (ONT.13798.5) as candidates affecting olecranon-type traits. - We next selected 11 DEGs that may affect olecranon- type traits, which are shown in Table 3. - Validation of expression patterns of candidate genes To confirm the accuracy of the ONT full-length tran- scriptome sequencing results and validate the expression patterns of the candidate genes, RT-PCR of the 11 selected genes was performed (Table 3). - The sequences of the gene-specific primers are shown in Table 4. - Complete correlations between RNA-Seq data and RT-PCR data were also identified in most of the tested genes (R = 1). - Therefore, the qRT-PCR data confirmed the reliability of the tran- scriptome data.. - There are obvious differences in the shape of round peach and olecranon honey peach (Fig. - Based on quality indicators, the round peach remark- ably differs from olecranon honey peach, particularly in the fruit type (Table 2). - A-3: Olecranon honey peach. - B-3: round peach. - type index, and the part of the fruit that resembles an eagle’s beak. - Table 3 Basic information on corresponding genes in the differential transcriptome. - In previous studies of peach fruit shape, some candidate genes for forming the flat peach have been identified, such as ppa003772m [5] and ppa025511m [6], but olecranon-type traits have not been evaluated. - In the current study, the olecranon-type trait was located close to the candidate genes identified by linkage analysis.. - ppa003772m and ppa025511m were not included within 1 Mb up- or down-stream of the main GWAS locus for olecranon-type traits (Supplemental Table 6). - PpCAD1 did not differ in the round peach and olecra- non honey peach according to ONT full-length sequencing.. - Furthermore, to better understand the formation of olecranon-type traits in peach at the molecular level, combined ONT full-length sequencing was performed to comprehensively analyze the transcriptome profile in the olecranon honey peach and round peach. - Based on their functional annotations and coding se- quences, 3 genes were identical among the 11 identi- fied candidate genes, indicating similar mechanisms in regulation of the olecranon-type trait and flat-type trait. - As a member of the B3 DNA binding superfamily, auxin-responsive factors can bind to auxin-responsive el- ements of induced genes and further regulate plant me- tabolism [26]. - The shape of the top of olecranon honey peach fruit resembles the beak of an eagle, which is popular among consumers. - No studies have been per- formed to examine the olecranon-type trait. - In this study, loci associated with the olecranon-type trait were examined by GWAS. - The gene loci related to the olecranon-type trait was bp in NC_034014.1.. - To further identify olecranon-type trait genes, we per- formed ONT full-length transcriptome sequencing of young fruits of 3 olecranon honey peaches and 3 round peaches. - The genes were within 1 Mb up- or down-stream of the main GWAS locus for olecranon-type traits. - These results provide a genetic basis for the olecranon-type trait and can be used to further evaluate the eagle beak shape of olecranon honey peach.. - A total of 456 peach accessions were collected in China for GWAS and are distributed in the North (315), Northwest (26), Qinghai-Tibet Region (3), South (83), Yangtze River Basin (22), and Yunnan-Guizhou Plateau (7) regions in China (Fig. - The tree was maintained in the same manner as other peach trees in the orchard.. - Ripening fruits were selected by experienced growers (fruit hanging time of approximately 75 days) and quickly transported to a well-ventilated, sterile labora- tory to count the olecranon type traits. - The values of the squared correlation coefficient (r 2 ) and significance of any LD detected between polymorphic sites were analyzed in all chromosomes with a 1000-kb window. - GWAS of the leaves was based on high-confidence SNPs (minor allele fre- quency >. - Mapped reads were further collapsed by using cDNA Cupcake package with coverage ≤ 0.85, identity ≤ 0.90, and merge sequences showing differences only in the 5′ exons.. - Validation of gene by quantitative real-time RT-PCR RNAs were isolated from the top of the olecranon honey peach and round peach. - Olecranon indi- cates whether the olecranon-type trait is present. - indicates the percent- age of bases at or above a given depth of coverage as a percentage of the total base number of the reference genome. - in- dicates the percentage of the sequenced data mapped to the reference genomes. - Statistical table of genes within 1Mb upstream and downstream of main genome-wide association study sites of olecranon type traits. - B-3: Round peach. - Statis- tical table of the number of annotated genes and transcripts. - Statistics of transcripts ex- pression in the round peach and the olecranon peach. - The nucleic acid sequence of the ONT gene is as follows.. - B-3: Round peach.. - The ordinate is the name of the KEGG metabolic pathway and abscess is the number of genes annotated to the pathway and its proportion to the total number of genes annotated.. - The original datasets analyzed in the current study are available in the NCBI Sequence Read Archive (SRA). - The sampling shall comply with the regulations of the people ’ s Republic of China on wildlife protection, and the samples were collected on private land with permission from the landowner. - Cytogenetical Studies in the Genus Citrus.. - A survey of the sorghum transcriptome using single- molecule long reads. - Distribution of SUN, OVATE, LC, and FAS in the tomato germplasm and the relationship to fruit shape diversity. - A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118
Xem thử không khả dụng, vui lòng xem tại trang nguồn hoặc xem
Tóm tắt