Gene structure of the AP2/ERF genefamily Gene structure analysisof Helianthus annuusAP2/ERFgenes further showed that the member withinthe subfamily possessed the similar exon-intron structures. As a whole, thenumber of exonregions ranged from 1 to 10, Soloist: 5 intron, RAV: 0-1 intron, AP2 and ANT: 4-9 introns, ERFandDREB: 0-1 intron. Of course, HaAP2/ERF-182 has 2 introns, HaAP2/ERF-215and HaAP2/ERF-217have 3 introns, and HaAP2/ERF-214including 4introns.
Most of the ERF subfamilygenes (86.29%) were observed to be intronless, which was consistent with theprevious study4. In contrast, the AP2 subfamily members containedmore intron thanERFs, which had at least fiveexons (FigureS2). The highlydiverse gene structure suggested that vast differentiation may occur during sunflowergenome formation and evolution. Duplication and divergence rate of the HaAP2/ERFgenes Gene duplication andsynteny analyses of AP2/ERFs between sunflower and other species (Arabidopsis,soybeanand rice) for the tandem and segmental duplication events of HaAP2/ERFgenes were investigated through 17chromosomes of sunflower (Fig. 5).
288 AP2/ERF gene clusters contained 40 pair tandemduplicated genes were identified, which located on chromosome 2, 3, 4, 5, 8, 9,12, and 13. Furthermore, 50 pairs of chromosomal segments duplication were alsofound (Fig. 6). To derive the origin and evolutionary relationships of AP2/ERFgenes, the comparative syntenic analysis between sunflower with other species (Arabidopsis,soybean andrice) was performed (Fig. 7a, b, c).
Most of HaAP2/ERF genesshowed syntenic bias towards particular chromosomes of Arabidopsis, soybeanandrice, which indicated that the chromosomal rearrangement events likeduplication and inversion may predominantly shape the distribution andorganization of AP2/ERF genes in these genomes.Thesubstitution rate of non-synonymous (Ka) versus synonymous (Ks) was aneffective measure to examine the positive selection pressure after duplication,wherein Ka/Ks =1 means neutral selection, Ka/Ks <1 stands for purifyingselection, and Ka/Ks >1 signifies accelerated evolution with positiveselection51. Furthermore, the divergence rate of the tandem andsegmental duplicated HaAP2/ERF genes were calculated to detect selectioninfluence (Table S3and S4).
The Ka/Ks ratio for tandem duplicated gene-pairs inHelianthus annuusAP2/ERF genes ranged from 0.05 to 1.33 with an averageof 0.42, whereas Ka/Ks for segmental duplicated gene-pairs ranged from 0.03 to1.81 with an average of 0.53.These results indicated that the duplicated HaAP2/ERFgenes were under strong purifying selection pressure and had gone throughsubstitution elimination and enormous selective constraint by natural selectionduring