Cloning and expression characteristics of PdMODD genes in Populus × euramericana

doi:10.12302/j.issn.1000-2006.202007015

JOURNAL OF NANJING FORESTRY UNIVERSITY ›› 2021, Vol. 45 ›› Issue (2): 43-50.doi: 10.12302/j.issn.1000-2006.202007015

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Cloning and expression characteristics of PdMODD genes in Populus × euramericana

ZHANG Tengqian¹(), ZHANG Weixi¹, DING Changjun¹, ZHANG Jing¹, HU Zanmin², SU Xiaohua¹^,^*()

1. State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing 100091, China
2. Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China

Received:2020-07-15 Accepted:2020-11-30 Online:2021-03-30 Published:2021-04-09
Contact: SU Xiaohua E-mail:tengqianzhang66@126.com;suxh@caf.ac.cn

Abstract

Abstract:

【Objective】To explore the function of PdMODD in the growth and development of Populus × euramericana, we analyzed their sequence characteristics, subcellular localization and expression patterns in response to salt and drought stress, which will provide a theoretical basis for revealing the molecular mechanisms of stress resistance and breeding new poplar varieties. 【Method】Based on the rice MODD amino acid sequence, Podel.08G114100, Podel.10G158900 and Podel.17G098500 were selected using BLAST in the genome of Populus deltoides WV94 v2.1. Using these gene sequences as references and Populus × euramericana ‘Bofeng 3 hao’ cDNA as template, three genes (PdMODD1,PdMODD2 and PdMODD3) were cloned. The structural characteristics of the PdMODD protein and its genetic relationship with its homologous proteins were analyzed using bioinformatics methods. The subcellular localization of PdMODD genes was analyzed using particle bombardment. The expression characteristics of PdMODD genes in different tissues and their response to different stresses were analyzed using qRT-PCR. 【Results】The open reading frames of PDMODD1-PDMODD3 genes were 1 065, 1 065 and 1 074 bp, encoding 354, 354 and 357 amino acids, respectively, all of which were unstable hydrophilic proteins located in the nucleus.PdMODD genes are all members of the NINJA family and contain three conserved domains, one of which is the transcriptional repression motif EAR. Phylogenetic analysis demonstrated that PdMODD1 protein was closely related to PtAFP2 (Populus trichocarpa) and located in the same branch as AtAFP1 (Arabidopsis thaliana) and MeAFP2 (Manihot esculenta). PdMODD2 protein had a close relationship with JcAFP2 (Jatropha curcas) and converged into a branch with AtAFP2; PdMODD3 was closely related to PtAFP3-1 and located in the same branch as QsAFP3 (Quercus suber) and AtAFP4. The subcellular localization results showed that PdMODD genes were localized in the nucleus. The results of tissue specificity analysis showed that PdMODD genes were highly expressed in the leaves and the least expressed in the roots. The expression patterns of stress response showed that under NaCl and PEG stress, PdMODD1, PdMODD2 and PdMODD3 were significantly upregulated in the root, stem and leaf tissues. 【Conclusion】PdMODD1-PdMODD3 are all members of the NINJA family, with the conserved transcriptional repression motif EAR and tissue specificity. They were highly expressed in the leaves, significantly induced by NaCl and PEG stress, and were in a branch with AtAFP1, AtAFP2 and AtAFP4, which had negative regulatory effects on ABA and salt stress. It is hypothesized that PdMODD1, PdMODD2 and PdMODD3 may play an important regulatory role in response to P. euramericana ‘Bofeng 3 hao’ salt and drought stress.

Key words: Populus × euramaricana ‘Bofeng 3 hao’, PdMODD, bioinformatics analysis, expression pattern analysis

CLC Number:

S785.5

ZHANG Tengqian, ZHANG Weixi, DING Changjun, ZHANG Jing, HU Zanmin, SU Xiaohua. Cloning and expression characteristics of PdMODD genes in Populus × euramericana[J]. JOURNAL OF NANJING FORESTRY UNIVERSITY, 2021, 45(2): 43-50.

Figures/Tables 8

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Table 2

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References 30

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用途 application	引物名称 primer name	上游引物序列(5'-3') forward primer sequence	下游引物序列(5'-3') reverse primer sequence
PdMODD基因克隆 cloning of PdMODD genes	PdMODD1	TAGAGAGGCAGCATCATGGGAG	TTACACAAAGGACGGACCAG
	PdMODD2	ATGGGAGAAGCAAACGCGAAT	TTATTCAAAGGACGGACAAGAAGG
	PdMODD3	GCTGGGATTTGAGGTTGTTGGT	TTAAAAAAAGGGAGAGGGGTGAACC
qRT-PCR	PdMODD1-RT	ATCAAGCAGTGGTGGTGAAG	CCTCCCTATATTTTCTGCAG
	PdMODD2-RT	GATCAAGCAGTGGTGGAGAAG	CCTCCCTCTATTTTCTGCAG
	PdMODD3-RT	TTTGAGAGCCAGCCACTT	CCATTCTCAGTGACTGTAAG
	actin	CATCCAGGCTGTCCTTTCCC	AACGAAGGATGGCGTGTGG
pBI121-PdMODD-GFP 融合表达载体构建 construction of pBI121- PdMODD-GFP fusion expression vector	PdMODD1-GFP	CTCTAGACTGGTACCCA TGGGAGAAGCAAACG	ACTAGTCAGTCGACCCGGTTC AAAGGACGGACAAG
	PdMODD2-GFP	CTCTAGACTGGTACCCATG GGAGAAACAAACG	ACTAGTCAGTCGACCCAAC ACAAAGGACGGACCAG
	PdMODD3-GFP	CTCTAGACTGGTACCCA TGGCACAAACAGAG	ACTAGTCAGTCGACCCGGA AAAAAGGGAGAGGGGTG

基因 gene	ORF 长度/bp ORF length	氨基酸数目 number of amino acid	分子质量/ ku relative molecular weight	理论等电点 pI	亚细胞定位 subcellular localization
PdMODD1	1 065	354	38.83	7.66	细胞核nucleus
PdMODD2	1 065	354	38.71	8.20	细胞核nucleus
PdMODD3	1 074	357	39.56	6.05	细胞核nucleus

Cloning and expression characteristics of PdMODD genes in Populus × euramericana

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