Cloning, sequencing, and expression analysis of 32 NAC transcription factors (MdNAC) in apple.

Background: NAC transcription factors play important roles in the regulation of plant growth, development, abiotic and biotic stress responses. The transcriptional level of s in different tissues and under various biotic and abiotic stress treatments was determined to provide a solid foundation for studying the function of s.

Methods: Thirty-two full-length cDNA sequences of s were isolated by homologous comparison and RT-PCR confirmation, and the obtained cDNA sequences and the deduced amino acid sequences were analyzed with bioinformatics methods. The prediction of subcellular locations of MdNAC proteins was performed using CELLO v.2.5, PSORT, and SoftBerry ProtComp 9.0. Expression levels of s were detected in 16 different tissues using an array. Expression patterns of s were detected in response to apple pathotype (AAAP) infection using RNA-seq, and the expression of s was analyzed under NaCl and mannitol treatments using RT-qPCR.

Results: The sequencing results produced 32 cDNAs (designated as , and with GenBank accession No. MG099861-MG099876, MG099891-MG099902, and MG099904-MG099907, respectively). Phylogenetic analysis revealed that MdNAC34 belonged to the ATAF group, MdNAC63 belonged to the AtNAC3 group, MdNAC24, MdNAC26-30, MdNAC32-33, MdNAC35, MdNAC37-39, MdNAC56-57, MdNAC59-62, MdNAC64-65, and MdNAC67-70 belonged to the NAM group, and MdNAC25, MdNAC36, MdNAC54-55, and MdNAC58 belonged to the VND group. Predictions of subcellular localization showed that MdNAC24-27, MdNAC29-30, MdNAC33-37, MdNAC39, MdNAC54-65, and MdNAC67-70 proteins were located in the nucleus, MdNAC28 proteins were located in the cytoplasm, MdNAC31-32 proteins were located in the nucleus and cytoplasm, and MdNAC38 proteins were located in the nucleus and plasma membrane. Array results indicated that 32 were expressed in all examined tissues at various expression levels. RNA-seq results showed that expression levels of , , , and were induced, but , and were down-regulated in response to AAAP infection. Under salt treatment, , , , , , , , and transcription levels were induced. Under mannitol treatment, and transcription levels were induced, but , , , , , , , , and were down-regulated. Taken together, the results indicated that the cloned genes were expressed constitutively in all examined tissues. These genes were up-regulated or down-regulated in response to AAAP infection and to salt or mannitol, which suggested they may be involved in the regulation of growth, development, and stress response in apple.