Cloning and expression analysis of the SpLEA1 gene of Selaginella pulvinata under drought stress

Abstract

Late embryogenesis abundant (LEA) is widely present in organisms and closely related to plant resistence, it can protect plant cells and reduce plant damage under drought stress. Selaginella pulvinata is a fern with the ability to survive drought stress, with a strong recovery ability under drought stress. To investigate the molecular mechanisms and expression characteristics of the SpLEA1 gene in drought-tolerant plants, we used the highly drought-tolerant plant S. pulvinata as experimental material and obtained the cDNA sequence of the SpLEA1 gene by RT-PCR based on the transcriptome sequencing results. The promoter sequence was obtained by the HiTail-PCR technique, and the sequence was analyzed by bioinformatics. qRT-PCR was used to analyze the expression pattern of the SpLEA1 gene under drought stress. The results were as follows: (1) The length of SpLEA1 was 476 bp, the open reading frame (ORF) was 279 bp, and it encoded 92 amino acids. The predicted molecular weight of the protein was 9 491.46 Da, and the isoelectric point was 5.45. The predicted protein structure analysis showed that the protein was hydrophilic. The protein contained ten phosphorylation sites, of which six serines, three tyrosines, and one threonine, respectively, and the predicted secondary structures showed that the protein was mainly composed of α-helix and random coil. (2) The conserved structural domain of the SpLEA1 protein was predicted to be Lea-5, derived from the LEA1 family. Based on the phylogenetic tree and genetic distanced matrix, the SpLEA1 was found to have high homology with Lea-5 protein from Cicer arietinum and Trifolium pratense. (3) Predictive analysis of cis-acting elements in promoter sequenced revealed that the SpLEA1 gene promoter contained five classes of hormone response elements and functional elements related to the drought stress response. The SpLEA1 gene was hypothesized to have multiple functions in the plant body and was closely related to drought stress response mechanisms. (4) SpLEA1 gene expression was up-regulated under natural dehydration treatment and peaked in 12 h. After rehydration treatment for 24 h, expression was significantly down-regulated. In summary, the SpLEA1 gene is likely to be involved in the regulation of drought stress response mechanisms in matted curly cypress. This results provide the reference for further studies on the function of the matted cypress SpLEA1 gene under drought stress and its expression regulation mechanism.

Keywords

• selaginella pulvinata
• splea1
• genetic cloning
• promoter cloning
• expression analysis