Genome-wide identification of CmCIPK family and its expression analysis under abiotic stress in melon

doi:10.3969/j.issn.1004-1524.2021.09.07

Acta Agriculturae Zhejiangensis ›› 2021, Vol. 33 ›› Issue (9): 1625-1639.DOI: 10.3969/j.issn.1004-1524.2021.09.07

• Horticultural Science • Previous Articles Next Articles

Genome-wide identification of CmCIPK family and its expression analysis under abiotic stress in melon

XIONG Xue(), ZHAO Lina, YANG Senlin, SAMIAH Arif, ZHANG Yidong^*()

College of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai 200240, China

Received:2020-11-10 Online:2021-09-25 Published:2021-10-09
Contact: ZHANG Yidong

Abstract

Abstract:

In order to explore the biological functions of CIPK(CBL-interacting protein kinase) gene family in melon, 18 melon CIPK family members were identified by the BLASTP based on the amino acid sequence of CIPK gene family of Arabidopsis thaliana, and their physico-chemical properties, chromosomal distribution, systematic evolution, gene structure, protein conserved motifs, cis-elements and gene expression patterns were analyzed. The results showed that gene length of CIPK family members was 1 499-8 499 bp, and they were unevenly distributed on 9 chromosomes of melon; According to the evolutionary relationship, they could be divided into five sub-families, which contain 26, 10, 25, 26 and 8 members, respectively. Among them, four pairs of CmCIPKs had fragmented duplication. Gene structure analysis showed that ten genes were intron-poor and eight genes were intron-rich. Protein conservation motif analysis found that the CmCIPK family was well-conserved, and all members contained typical features of the CIPK family: the activation loop in the N-terminal kinase domain and the NAF/FISL domain in the C-terminal regulatory domain. The presence of multiple cis-elements related to plant hormones and stress in the 2 000 bp upstream sequence of their genes suggested that there might be multiple transcriptional regulation. Transcriptome analysis found that the tissue expression levels of CmCIPKs from high to low were: leaf, root, male flower, female flower and fruit. The CmCIPK1-like and CmCIPK12-like genes were selected to verify their tissue expression patterns, which showed the highest expression levels in leaves and male flowers, respectively. After different adversity treatments, the results showed that the two CmCIPK genes were induced by NaCl and abscisic acid (ABA), and they returned to the initial expression level after a short period of up-regulation or down-regulation in drought treatment. The results indicated that CmCIPK1-like and CmCIPK12-like might play important roles in ABA and abiotic stress, which could provide references for future CmCIPK functional studies.

Key words: melon (Cucumis melo L.), CIPK family, genome-wide analysis, expression patern, abiotic stress

CLC Number:

S652.9
Q78

XIONG Xue, ZHAO Lina, YANG Senlin, SAMIAH Arif, ZHANG Yidong. Genome-wide identification of CmCIPK family and its expression analysis under abiotic stress in melon[J]. Acta Agriculturae Zhejiangensis, 2021, 33(9): 1625-1639.

Figures/Tables 13

Table 1 Primers for real-time PCR

基因Gene	上游引物序列Forward primer(5'→ 3')	下游引物序列Reverse primer(5'→ 3')
CmActin	ACATCTGCTGGAAGGTGCTT	CCCTGGTATTGCAGACAGGA
CmCIPK1-like	TCCATGCATGAAGTGCCATCT	ACAACATCCTCCATCCGCTT
CmCIPK12-like	GAGATCTCAATCCTCCGCCG	CGCGATGATAAACACCACGC

Table 2 Basic information of CmCIPK gene family

基因名称 Gene name	基因号 Gene ID	基因长度 Gene length/bp	编码蛋白氨基酸数量 Number of amino acids encoding protein/aa	分子量 Relative molecular weight/ku	等电点 PI	亲水性平均值 Grand average of hydropathicity	不稳定系数 Instability index
CmCIPK1	MELO3C006758.2	3 023	442	49.4	5.60	-0.373	40.19
CmCIPK1-like	MELO3C021231.2	7 022	459	52.8	7.26	-0.235	42.96
CmCIPK2	MELO3C026055.2	3 380	468	53.1	8.73	-0.371	28.26
CmCIPK3	MELO3C014700.2	8 247	444	50.5	6.91	-0.399	34.34
CmCIPK5-like	MELO3C016967.2	1 793	443	50.2	8.60	-0.297	39.42
CmCIPK6-like	MELO3C010234.2	1 948	430	48.5	9.07	-0.381	37.43
CmCIPK7-like	MELO3C014269.2	1 672	430	47.8	9.05	-0.249	40.48
CmCIPK8	MELO3C011108.2	6 254	446	50.7	7.58	-0.280	41.25
CmCIPK9	MELO3C005987.2	4 322	441	50.1	8.51	-0.449	33.75
CmCIPK10-like	MELO3C026873.2	2 410	471	53.3	9.12	-0.451	38.89
CmCIPK11	MELO3C027266.2	1 652	425	48.2	8.48	-0.361	42.96
CmCIPK12-like	MELO3C007208.2	2 109	467	53.0	8.26	-0.334	41.95
CmCIPK14	MELO3C026058.2	1 499	433	49.1	6.08	-0.242	40.35
CmCIPK20	MELO3C002661.2	2 402	463	52.4	9.25	-0.422	33.62
CmCIPK23	MELO3C007110.2	5 130	455	51.2	8.74	-0.316	33.88
CmCIPK23-like	MELO3C002766.2	8 499	465	52.2	9.04	-0.444	35.14
CmCIPK24	MELO3C010334.2	6 644	437	49.7	7.64	-0.297	47.75
CmCIPK25-like	MELO3C026741.2	1 750	445	50.2	8.81	-0.227	34.58

Fig.1 Location of CmCIPK on the chromosomes of melon and collinearity analysis A, Specific distribution of members of the CmCIPKs on each chromosome. The size of the chromosome was represented by its relative length. The chromosomes without the CmCIPK(1st, 9th and 10th) were not shown in the figure. B, Collinearity analysis of CmCIPK. The black line represented the gene pair in the CmCIPK family where fragment replication occurred, and the gray line represented all the gene pairs in the melon genome where fragment replication occurred.

Fig.2 Phylogenetic analysis of CIPK proteins in melon, Arabidopsis, rice and cucumber 18 CmCIPK proteins (diamond), 26 AtCIPK proteins (circle), 33 OsCIPK proteins( triangle) and 18 CsCIPK proteins (square) were used to construct the neighbor tree by MEGA 7.0. The five groups were marked as Ⅰ, Ⅱ, Ⅲ, Ⅳ, Ⅴ.

Fig.3 Exon-intron structure of CmCIPKs Use TBtools software to draw gene structure, and arrange according to phylogenetic tree. The yellow boxes represented exons, the green boxes represented UTRs, and the lines represented introns.

Fig.4 Motif analysis of CmCIPK proteins A, Use MEME to identify conserved motifs and arrange them according to the phylogenetic tree. Each motif was represented by a colored box, and its name was given on the right. B, Annotation of motif sequence encoding known functional domain. The total height of each stack indicated the sequence conservation at that position, and the height of the letters in each stack represented the degree of conservation of amino acids in the motif.

Fig.5 Promoter cis-elements analysis of CmCIPK gene family ABRE, ABA-responsive element; TGA-element, auxin-responsive element; TC-rich repeats, defense and stress response element; LTR, low-temperature-responsive element; TCA-element, salicylic acid response element; W-box, defense and stress response element; MBS, drought stress response element; CGTCA-motif, Methyl jasmonate-responsive element.

Fig.6 Transcription abundance of CmCIPK in different tissues of melon Use Graphpad 8.2.1 to draw the heat map, the scale represented the relative size of the log2FPKM value, green represented low transcription abundance, and red represented high transcription abundance.

Fig.7 Expression level of CmCIPK1-like and CmCIPK12-like in different tissues of melon Different letters indicated significant differences (P<0.05). The same as below.

Fig.8 Relative expression level of CmCIPK1-like and CmCIPK12-like in leaves of melon under 100 μmol·L-1 ABA treatment

Fig.9 Relative expression level of CmCIPK1-like and CmCIPK12-like in roots, stems and leaves of melon under NaCl treatment for 5 days

Fig.10 Relative expression level of CmCIPK1-like and CmCIPK12-like in roots, stems and leaves of melon under 200 mmol·L-1 NaCl treatment

Fig.11 Relative expression level of CmCIPK1-like and CmCIPK12-like in roots, stems and leaves of melon under 15% PEG treatment

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Genome-wide identification of CmCIPK family and its expression analysis under abiotic stress in melon

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