ABSTRACT: Purple carrots can accumulate large quantities of anthocyanins in their roots. Depending on the genetic background, anthocyanin pigmentation can be expressed in the entire root, or it can display tissue specific-patterns, confined to the root phloem or xylem tissues. Within the phloem, the tissue usually contributing most of the overall anthocyanin concentration in the carrot root, purple pigmentation can be found in the outer phloem (OP) (also called cortex) and inner phloem (IP) tissues, or it can be confined exclusively to the OP. The latter is a fairly-common phenotype in many purple carrot cultivars. In this work, the genetic control underlying tissue-specific anthocyanin pigmentation in the carrot root OP and IP tissues was investigated by means of linkage mapping, transcriptome (RNA-seq), phylogenetic, and gene expression (RT-qPCR) analyses in two genetic backgrounds; an F2 mapping population (3242) and the inbred line B7262. Genetic mapping of the ‘root outer phloem anthocyanin pigmentation’ (ROPAP) and inner phloem pigmentation (RIPAP) revealed co-localization of ROPAP with the P1 and P3 genomic regions previously known to condition pigmentation in different genetic stocks, whereas RIPAP co-localized with P3 only. Transcriptome analysis of purple OP (POP) vs. non-purple IP (NPIP) tissues, along with linkage and phylogenetic data, allowed an initial identification of 28 candidate genes, 19 of which were further evaluated by RT-qPCR in independent root samples of 3242 and B7262, revealing 15 genes consistently upregulated in the POP in both genetic backgrounds, and two genes upregulated in the POP in specific backgrounds. These include seven transcription factors (4 MYBs, 1 bHLH, 1 MADS-box, 1 ERF), seven anthocyanin structural genes, and two genes involved in cellular transport. Altogether, our results point at DcMYB7, DcMYB113, and a MADS-box (DCAR_010757) as the main candidate genes conditioning ROPAP in 3242, whereas DcMYB7 and MADS-box condition RIPAP in this background. In 7262, which roots present purple pigmentation only in the outer phloem, DcMYB113 conditions ROPAP
