Garlic Circadian Clock Genes AsRVE1 and AsRVE2 and Their Expression Analysis Under Osmotic Stress

doi:10.16420/j.issn.0513-353x.2020-0961

Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (9): 1706-1716.doi: 10.16420/j.issn.0513-353x.2020-0961

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Garlic Circadian Clock Genes AsRVE1 and AsRVE2 and Their Expression Analysis Under Osmotic Stress

BIAN Shicun¹, LU Yani¹, XU Wujun¹, CHEN Boqing², WANG Guanglong²^,^*(), XIONG Aisheng³^,^*()

¹Faculty of Applied Technology,Huaiyin Institute of Technology,Huaian,Jiangsu 223003,China
²School of Life Science and Food Engineering,Huaiyin Institute of Technology,Huaian,Jiangsu 223003,China
³College of Horticulture,State Key Laboratory of Crop Genetics and Germplasm Enhancement,Ministry of Agriculture and Rural Affairs Key Laboratory of Biology and Germplasm Enhancement of Horticultural Crops in East China,Nanjing Agricultural University,Nanjing 210095,China

Received:2021-05-13 Revised:2021-07-31 Online:2021-09-25 Published:2021-09-30
Contact: WANG Guanglong,XIONG Aisheng E-mail:guanglongwang@hyit.edu.cn;xiongaisheng@njau.edu.cn

Abstract

Abstract:

In order to understand the sequence characteristics of circadian clock-related genes REVEILLEs（RVEs）in garlic and their roles in osmotic stress response,AsRVE1 and AsRVE2 genes were isolated from garlic,and their expression patterns under salt stress and simulated drought stress were analyzed by qRT-PCR. Sequence analysis showed that the open reading frames of AsRVE1 and AsRVE2 genes were 1 050 and 975 bp in length,encoding 349 and 324 amino acids,respectively. In terms of evolutionary relationship,AsRVE1 and AsRVE2 were relatively close to ZmRVE2 in corn and AcRVE2 in pineapple. The RVE amino acid sequences of different plants had low homology,but showed high consistency in the conserved domain at the N-terminus. Both AsRVE1 and AsRVE2 genes can respond to changes in circadian rhythms,and their expression can be detected in different tissues;the expression of AsRVE1 in different tissues was not obviously different,the expression of AsRVE2 in the roots was relatively high. Both drought stress and salt stress induced the altered expression of AsRVE1 and AsRVE2 genes in different tissues. These results indicated that AsRVE1 and AsRVE2 genes may be involved in drought- and salt-stress response in garlic. The AsRVE1 and AsRVE2 genes cloned in this study laid a foundation for further identification of the biological functions and regulatory mechanisms of RVE.

Key words: garlic, circadian rhythm, RVE transcription factor, osmotic stress, gene cloning, expression analysis

CLC Number:

S633.4

BIAN Shicun, LU Yani, XU Wujun, CHEN Boqing, WANG Guanglong, XIONG Aisheng. Garlic Circadian Clock Genes AsRVE1 and AsRVE2 and Their Expression Analysis Under Osmotic Stress[J]. Acta Horticulturae Sinica, 2021, 48(9): 1706-1716.

Figures/Tables 10

Table 1 Primer sequences used for gene cloning and quantitative real time fluorescence PCR

用途 Use	基因名称 Gene name	引物（上游/下游） Primer（Forward/Reverse）
克隆 Gene cloning	AsRVE1	ATGACAAGTGATCCTTTATACTTGG/TCATCTCTTGTATGGAAGAAATCCT
克隆 Gene cloning	AsRVE2	ATGCAGATCAGTAGTACATCACCTG/TCACCTCCTGTACGGAACAAATCCC
荧光定量PCR Quantitative real time fluorescence PCR	AsRVE1	TCAGTGGATGTGGAGAACAATACCT/TGATGAGTAAGGAGGCAACAGATGA
	AsRVE2	ACAGCGTGAGAGGTGGACTGAT/GAGATGCAGACGGCGAACCAA
	AsSAND	GCGTCAACGAATGTTCCAATTACCA/TCTCTTCAGTCTCAACTTCATCAGCAT

Fig. 1 AsRVE1 and AsRVE2 genes amplified from garlic by RT-PCR

Fig. 2 Nucleotide sequences and amino acid sequences of garlic AsRVE1（left）and AsRVE2（right）genes

Fig. 3 Phylogenetic tree of RVE amino acid sequences from garlic and other plants

Fig. 4 Alignment of RVE amino acid sequences from garlic and other plant As：Allium sativum;At：Arabidopsis thaliana;Ls：Lactuca sativa;Zm：Zea mays;Rs：Raphanus sativus;Vr：Vigna radiata;Ac：Ananas comosus. * indicates DNA binding sites.

Fig. 5 Prediction of secondary structure of AsRVE1 and AsRVE2 of garlic Blue,red,green,and purple respectively represent alpha helix,extended strand,beta turn,and random coil.

Fig. 6 Protein-protein interaction network of garlic AsRVE1 and AsRVE2

Fig. 7 Response of garlic AsRVE1 and AsRVE2 to light period Different lowercase letters represent significant differences at 0.05 level.

Fig. 8 Expression level of garlic AsRVE1 and AsRVE2 under drought stress Different lowercase letters in the same tissue represent significant differences at 0.05 level.

Fig. 9 Expression level of garlic AsRVE1 and AsRVE2 under salt stress Different lowercase letters in the same tissue represent significant differences at 0.05 level.

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Garlic Circadian Clock Genes AsRVE1 and AsRVE2 and Their Expression Analysis Under Osmotic Stress

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