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Molecular cloning and characterization of NPR1 gene from Arachis hypogaea

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Abstract

The NPR1 gene was an important regulator for a plant disease resistance. The cDNA of NPR1 gene was cloned from peanut cultivar Ri Hua 1 by rapid amplification of cDNA ends-polymerase chain reaction (RACE-PCR). The full length cDNA of Arachis hypogaea NPR1 consisted of 2,078 base pairs with a 1,446 bp open-reading frame encoding 481 amino acids. The predicted NPR1 contained the highly conserved functional domains (BTB/POZ domain from M1 to D116), protein–protein interaction domains (three ankyrin repeats from K158 to L186; N187 to L217 and R221 to D250) and one NPR1-like domain (C262 to S469). The DNA sequence of the NPR1 gene was 2,332 or 2,223 bp. Both two sequences contained three introns and four exons. The NPR1 transcripts were expressed mainly in roots and leaves, while fewer signals were detected in the stems. Amount of the NPR1 transcript was significantly increased 1 h after salicylic acid challenge and was eventually 5.3 times greater than that in the control group. Both the DNA sequence and the coding sequence were obtained from eight cultivars and nine wild species of Arachis. Maximum likelihood analyses of d N/d S ratios for 25 sequences from different species showed that different selection pressures may have acted on different branches.

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Abbreviations

bp:

Base pair

BW:

Bacterial wilt

df:

Degrees of freedom

EST:

Expressed sequence tag

LRT:

Likelihood ratio test

NPR1 :

Nonexpressor of pathogenesis-related genes1

PR genes:

Pathogenesis-related genes

qRT-PCR:

Quantitative real-time reverse transcription polymerase chain reaction

RT-PCR:

Reverse transcription polymerase chain reaction

SA:

Salicylic acid

SAR:

Systemic acquired resistance

UTR:

Untranslated region

TGA:

Leucine zipper transcription factor TGA

TUA:

Alpha-tubulin

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Acknowledgments

The authors are grateful to all the laboratory members for continuous technical advice and helpful discussion. This research was supported from China Agricultural Research System (CARS-14), Qingdao Science and Technology plan Basic research Project (12-1-4-11-(1)-jch).

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Authors and Affiliations

  1. Shandong Peanut Research Institute (SPRI), 126 Fushan Rd., Qingdao, 266100, People’s Republic of China

    Qi Wu, Xiu Zhen Wang, Yue Yi Tang, Hong Tao Yu, Yu Fei Ding, Chuan De Yang, Feng Gao Cui, Jian Cheng Zhang & Chuan Tang Wang

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  1. Qi Wu
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  7. Feng Gao Cui
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Correspondence to Chuan Tang Wang.

Electronic supplementary material

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Appendix S1 Fig. S1 Multiple alignment of NPR1 sequences from A. hypogaea cultivar, Ri Hua 1 (DOC 101 kb)

11033_2014_3395_MOESM2_ESM.doc

Appendix S2 Fig. S2 Ratios of d N/d S for NPR1 for the indicated branches. The ω values of each branch were the result of the free ratio model. A ω < 1 indicates that a purifying selection or a relaxed selective constraint has acted along that lineage. ‘Inf’ indicates infinity. Nodes marked with ‘1’ ‘2’ or ‘3’ mean that the clades have been tested by the branch site model A. The three clades represent the lineage of genus Arachis and the two dicotyledon clades, respectively. (DOC 91 kb)

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Wu, Q., Wang, X.Z., Tang, Y.Y. et al. Molecular cloning and characterization of NPR1 gene from Arachis hypogaea . Mol Biol Rep 41, 5247–5256 (2014). https://doi.org/10.1007/s11033-014-3395-z

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  • DOI: https://doi.org/10.1007/s11033-014-3395-z

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