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Background Mitogen-activated protein kinase (MAPK) cascades play vital roles in signal transduction in response to a wide range of biotic and abiotic stresses. In a previous study, we identified ten ZjMAPKs and five ZjMAPKKs in the Chinese jujube genome. We found that some members of ZjMAPKs and ZjMAPKKs may play key roles in the plant’s response to phytoplasma infection. However, how these ZjMAPKKs are modulated by ZjMAPKKKs during the response process has not been elucidated. Little information is available regarding MAPKKKs in Chinese jujube. Results A total of 56 ZjMAPKKKs were identified in the jujube genome. All of these kinases contain the key S-TKc (serine/threonine protein kinase) domain, which is distributed among all 12 chromosomes. Phylogenetic analyses show that these ZjMAPKKKs can be classified into two subfamilies. Specifically, 41 ZjMAPKKKs belong to the Raf subfamily, and 15 belong to the MEKK subfamily. In addition, the ZjMAPKKKs in each subfamily share the same conserved motifs and gene structures. Only one pair of ZjMAPKKKs (15/16, on chromosome 5) was found to be tandemly duplicated. Using qPCR, the expression profiles of these MAPKKKs were investigated in response to infection with phytoplasma. In the three main infected tissues (witches’ broom leaves, phyllody leaves, and apparently normal leaves), ZjMAPKKK26 and − 45 were significantly upregulated, and ZjMAPKKK3, − 43 and − 50 were significantly downregulated. ZjMAPKKK4, − 10, − 25 and − 44 were significantly and highly induced in sterile cultivated tissues infected by phytoplasma, while ZjMAPKKK6, − 7, − 17, − 18, − 30, − 34, − 35, − 37, − 40, − 41, − 43, − 46, − 52 and − 53 were significantly downregulated. Conclusions For the first time, we present an identification and classification analysis of ZjMAPKKKs. Some ZjMAPKKK genes may play key roles in the response to phytoplasma infection. This study provides an initial understanding of the mechanisms through which ZjMAPKKKs are involved in the response of Chinese jujube to phytoplasma infection.

Additional file 1: Figure S1. Protein sequences of MAPKKKs from Ziziphus jujuba Mill. and Arabidopsis thaliana.

Additional file 3: Figure S2. Healthy and diseased in vitro plantlets. A: Healthy plantlets; B: Diseased plantlets.

Additional file 4: Figure S3. Tissues showing different JWB disease symptoms. A: Witches’ broom leaves; B: Phyllody leaves; C: Apparently normal leaves; D: Healthy leaves. A, B and C were used as a test group collected from diseased trees. D was used as a control collected from healthy trees.

Additional file 7: Figure S5. CDS sequences of ZjMAPKKKs.

Additional file 5: Figure S4. Determination of phytoplasma in the sieve element in jujube petiole phloem by using 4′,6-diamidino-2-phenylindole (DAPI). A, No fluorescent spots were observed in the sieve element (SE) of healthy plantlets. B, The fluorescent spots formed a large, bright circle in the sieve element (SE) of the diseased plantlets. The numbers and sizes of the fluorescent spots indicate the number of phytoplasmas. Bar = 100 μm.

Backgrounds Mitogen activated protein kinase (MAPK) cascades play vital roles in signal transduction in response to various biotic and abiotic stresses. In the previous study we have identified 10 ZjMAPKs and 5 ZjMAPKKs in Chinese jujube genome and found some crucial members of ZjMAPKs and ZjMAPKKs might function importantly in the process of phytoplasma infection. But how these ZjMAPKKs were modulated by ZjMAPKKKs during this process is still elusive and little information is known about the MAPKKKs in Chinese jujube. Results In the current study, 56 ZjMAPKKKs were identified in the jujube genome and all of them contain the key S-TKc (serine/threonine protein kinase) domain which distributed in all 12 chromosomes. Phylogenetic analysis showed that these ZjMAPKKKs could be classified into two subfamilies, of which 41 belonged to Raf, and 15 to MEKK subfamily. In addition, the ZjMAPKKKs in each subfamily share the same conserved motifs and gene structures, one pair of ZjMAPKKKs (15/16) was the only tandem duplication event on Chromosome 5. Furthermore, the expression profiles of these MAPKKKs in response to phytoplasma disease were investigated by qPCR. In the three main infected tissues (witches’ broom leaves, phyllody leaves, apparent normal leaves), the ZjMAPKKK26 and 45 were significantly up regulated and the ZjMAPKKK3, 43 and 50 were down regulated. While the ZjMAPKKK4, 10, 25 and 44 were significant highly induced in the sterile cultivated tissues infected by phytoplasma, and the ZjMAPKKK7, 30, 35, 37, 40, 41, 43 and 46 were significantly down regulated. Conclusions The identification and classification analysis of ZjMAPKKKs was firstly reported and some key individual ZjMAPKKKs genes might play essential roles in response to phytoplasma infection. This could provide initial understanding for the mechanism that how the ZjMAPKKKs were involved in jujube - phytoplasma infection.