Cloning and Analyzing of <i>AP</i>3-3 and Its Promoter from <i>Dendrobium officinale</i>

WANG Yiqin; SUN Bo; HE Ling; SHI Kaihui; HUANG Xin; WANG Dongmei; CHEN Yu; ZANG Rui; HE Fengmei

doi:10.19303/j.issn.1008-0384.2022.005.005

Volume 37 Issue 5

May 2022

Turn off MathJax

Article Contents

Article Navigation > Fujian Journal of Agricultural Sciences > 2022 > 37(5): 585-591

WANG Y Q, SUN B, HE L, et al. Cloning and Analyzing of AP3-3 and Its Promoter from Dendrobium officinale [J]. Fujian Journal of Agricultural Sciences，2022，37（5）：585−591 doi: 10.19303/j.issn.1008-0384.2022.005.005

Citation:

WANG Y Q, SUN B, HE L, et al. Cloning and Analyzing of AP3-3 and Its Promoter from Dendrobium officinale [J]. Fujian Journal of Agricultural Sciences，2022，37（5）：585−591 doi: 10.19303/j.issn.1008-0384.2022.005.005

Citation:

PDF( 1543 KB)

Cloning and Analyzing of AP3-3 and Its Promoter from Dendrobium officinale

doi: 10.19303/j.issn.1008-0384.2022.005.005

College of Landscape Architecture and Horticulture, Yunnan Agricultural University, Kunming, Yunnan　650201, China

Received Date: 2022-02-21
Rev Recd Date: 2022-04-15

Available Online: 2022-06-20

Publish Date: 2022-05-28

Abstract

Abstract

Objective AP3-3 of class B gene of MADS box family that involves in the formation of perianth and labellum of Dendrobium officinale was cloned to study the biological function, and the promoter analyzed to decipher the regulation mechanism. Method Sequences of AP3-3 and promoter of D. officinale were cloned by RT-PCR and conventional PCR, and bioinformatics analyzed. A fusion expression vector of promoter-deleted fragments and GUS gene was constructed. Agrobacterium tumefaciens-mediated transformation was used to transform protocorm of D. officinale for the transient expression. Result The cDNA length of DoAP3-3 was 675 bp with an encoded formula of C₁₁₂₉H₁₈₀₃N₃₃₃O₃₄₇S₁₂, a molecular weight of 25.98 kDa, a PI of 8.71, an instability index of 40.14, and GRAVY of −0.823. There was no transmembrane region detected in the protein. The predicted score of subcellular localization was 87.0% in nucleus, 8.7% in mitochondria, and 4.3% in cytoplasm. The 1885 bp promoter fragment had a cis acting element containing a significant number of photo-responsive elements among others. The 3 promoter fragments could drive GUS with an order of expression intensity of −885–0 bp＞−1 604–0 bp＞−750–0 bp. Conclusion The predicted DoAP3-3 was an alkali, hydrophilic, and unstable protein with no transmembrane domain and a subcellular localization in the nucleus. The DoAP3-3 promoter might be regulated by light, plant hormones, MYB transcription protein, etc., and exhibited activities that increased with decreasing deletion length.
- Dendrobium officinale,
- AP3-3,
- cDNA,
- promoter,
- cloning,
- sequence analysis

FullText(HTML)

References(18)

References

[1]	LITT A, KRAMER E M. The ABC model and the diversification of floral organ identity [J]. Seminars in Cell & Developmental Biology, 2010, 21(1): 129−137.
[2]	王莹, 穆艳霞, 王锦. MADS-box基因家族调控植物花器官发育研究进展 [J]. 浙江农业学报, 2021, 33(6):1149−1158. WANG Y, MU Y X, WANG J. Research progress of floral development regulation by MADS-box gene family [J]. Acta Agriculturae Zhejiangensis, 2021, 33(6): 1149−1158.(in Chinese)
[3]	HUANG F Y, ZHANG Y H, HOU X L. BcAP3, a MADS box gene, controls stamen development and male sterility in Pak-choi (Brassica rapa ssp. chinensis) [J]. Gene, 2020, 747: 144698. doi: 10.1016/j.gene.2020.144698
[4]	IRISH V. The ABC model of floral development [J]. Current Biology, 2017, 27(17): R887−R890. doi: 10.1016/j.cub.2017.03.045
[5]	PAN Z J, CHENG C C, TSAI W C, et al. The duplicated B-class MADS-box genes display dualistic characters in orchid floral organ identity and growth [J]. Plant and Cell Physiology, 2011, 52(9): 1515−1531. doi: 10.1093/pcp/pcr092
[6]	DENG M H, LV J H, WANG Z R, et al. Two promoter regions confer heat-induced activation of SlDREBA4 in Solanum lycopersicum [J]. Biochemical and Biophysical Research Communications, 2020, 524(3): 689−695. doi: 10.1016/j.bbrc.2020.01.153
[7]	王树军, 刘保华, 孙进华, 等. 荔枝多酚氧化酶基因启动子克隆与功能分析 [J]. 果树学报, 2015, 32(3):427−433,524. WANG S J, LIU B H, SUN J H, et al. Cloning and function analysis of the promoter of PPO gene from Litchi(Litchi chinensis Sonn.) [J]. Journal of Fruit Science, 2015, 32(3): 427−433,524.(in Chinese)
[8]	HOU J J, JIANG P P, QI S M, et al. Isolation and functional validation of salinity and osmotic stress inducible promoter from the maize type-II H+-pyrophosphatase gene by deletion analysis in transgenic tobacco plants [J]. PLoS One, 2016, 11(4): e0154041. doi: 10.1371/journal.pone.0154041
[9]	XIN S, TAO C C, LI H B. Cloning and functional analysis of the promoter of an ascorbate oxidase gene from Gossypium hirsutum [J]. PLoS One, 2016, 11(9): e0161695. doi: 10.1371/journal.pone.0161695
[10]	李泽卿. 二球悬铃木花发育基因PaAP3、PaPI和PaSTK启动子的克隆、功能分析及其在不育中的应用[D]. 武汉: 华中农业大学, 2017. LI Z Q. Cloning and functional analysis of the promoters of flower development genes PaAP3, PaPI and PaSTK in London plane tree and their application in sterility[D]. Wuhan: Huazhong Agricultural University, 2017. (in Chinese)
[11]	ROMBAUTS S, DÉHAIS P, VAN MONTAGU M, et al. PlantCARE, a plant Cis-acting regulatory element database [J]. Nucleic Acids Research, 1999, 27(1): 295−296. doi: 10.1093/nar/27.1.295
[12]	张婷, 邢妮, 王超, 等. 小花草玉梅正常和自然变异植株的AP3-3基因研究 [J]. 西北植物学报, 2016, 36(2):231−240. ZHANG T, XING N, WANG C, et al. Cloning and sequence analysis of AP3-3 gene in normal plant and natural variant from Anemone rivularis var. flore-minore [J]. Acta Botanica Boreali-Occidentalia Sinica, 2016, 36(2): 231−240.(in Chinese)
[13]	JING D L, CHEN W W, SHI M, et al. Ectopic expression of an Eriobotrya japonica APETALA3 ortholog rescues the petal and stamen identities in Arabidopsis ap3-3 mutant [J]. Biochemical and Biophysical Research Communications, 2020, 523(1): 33−38. doi: 10.1016/j.bbrc.2019.11.177
[14]	李淑娴. 墨兰成花机理及花期调控技术研究[D]. 福州: 福建农林大学, 2016. LI S X. Mechanism of flower development and early flowering technique of Cymbidium sinense[D]. Fuzhou: Fujian Agriculture and Forestry University, 2016. (in Chinese)
[15]	龚湉. 寒兰成花机理及花期调控研究[D]. 福州: 福建农林大学, 2015. GONG T. Mechanism of floral formation of Cymbidium kanran and flowering regulation[D]. Fuzhou: Fujian Agriculture and Forestry University, 2015. (in Chinese)
[16]	MUNDY J, YAMAGUCHI-SHINOZAKI K, CHUA N H. Nuclear proteins bind conserved elements in the abscisic acid-responsive promoter of a rice rab gene [J]. PNAS, 1990, 87(4): 1406−1410. doi: 10.1073/pnas.87.4.1406
[17]	李艳林, SHAHID IQBAL, 侍婷, 等. 梅PmARF17克隆及其在花发育中与内源激素的调控模式 [J]. 中国农业科学, 2021, 54(13):2843−2857. doi: 10.3864/j.issn.0578-1752.2021.13.013 LI Y L, IQBAL S, SHI T, et al. Isolation of PmARF17 and its regulation pattern of endogenous hormones during flower development in Prunus mume [J]. Scientia Agricultura Sinica, 2021, 54(13): 2843−2857.(in Chinese) doi: 10.3864/j.issn.0578-1752.2021.13.013
[18]	POBORILOVA Z, PLCHOVA H, CEROVSKA N, et al. Transient protein expression in tobacco BY-2 plant cell packs using single and multi-cassette replicating vectors [J]. Plant Cell Reports, 2020, 39(9): 1115−1127. doi: 10.1007/s00299-020-02544-w