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| Cloning and Promoter Function Analysis of the Anthocyanins Synthase Gene (LsANS) in Lycoris sprengeri |
| XUE Hui-Min, ZHOU Yang-Li, GAO Yan-Hui* |
| State Key Laboratory of Subtropical Silviculture, Zhejiang A&F University, Hangzhou 311300, China |
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Abstract Lycoris sprengeri is multicolor special flowera with red and blue petals in Lycoris, which flower color formation mechanism is complex, and anthocyanin synthase (ANS) is a key enzyme at the end of plant anthocyanin synthesis pathway. In this study, molecular biological methods were used to analyze the functions of LsANS cDNA and promoter. The results showed, The total length of LsANS cDNA was 1 401 bp (GenBank No. MT664240), ORF 1 068 bp, encoding 355 amino acids; The phylogenetic tree analysis showed that the LsANS had highest homology with ANS gene of L. radiate, which was 98.5%. qRT-PCR showed that the expression of LsANS was the highest in clone H4 and the lowest in clone H1; The expression was the highest at large bud stage and the lowest at full-bloom stage; The consequence of HPLC showed that in different developmental stages, the content of cyanidin-3-O-glucoside and pelargonin-3-O-glucoside in the bud stage was higher than that in other stages; in different clones, pelargonin-3-O-glucose accounted for the highest proportion of anthocyanins measured in clone H4, and the lowest proportion in clone H1; It was speculated that LsANS played an important role in the anthocyanin formation pathway. A 2 114 bp long LsANS promoter sequence was cloned, which contained MYB transcription factor binding site and cis acting elements involved in light, hormone and stress. The PBI121 LsANSpro::GUS fusion expression vector was constructed and transiently transformed into Arabidopsis thaliana, which showed that LsANSpro has the function of promoting downstream genes. By GUS staining assay, promoter activity was detected in the whole roots, leaves, and flowers . The research will provide a theoretical basis for further research the role and mechanism of ANS gene in the flower color formation process of Lycoris.
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Received: 23 September 2021
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Corresponding Authors:
*gaoyanhui408@126.com
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