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Evaluation of Plant-Derived Promoters for Constitutive and Tissue-Specific Gene Expression in Potato
oleh: Dmitry Miroshnichenko, Aleksey Firsov, Vadim Timerbaev, Oleg Kozlov, Anna Klementyeva, Lyubov Shaloiko, Sergey Dolgov
Format: | Article |
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Diterbitkan: | MDPI AG 2020-11-01 |
Deskripsi
Various plant-derived promoters can be used to regulate ectopic gene expression in potato. In the present study, four promoters derived from the potato genome have been characterized by the expression of identical cassettes carrying the fusion with the reporter β-glucuronidase (<i>gusA</i>) gene. The strengths of <i>StUbi</i>, <i>StGBSS</i>, <i>StPat</i>, and <i>StLhca3</i> promoters were compared with the conventional constitutive CaMV <i>35S</i> promoter in various organs (leaves, stems, roots, and tubers) of greenhouse-grown plants. The final amount of gene product was determined at the post-transcriptional level using histochemical analysis, fluorometric measurements, and Western blot analysis. The promoter strength comparison demonstrated that the <i>StUbi</i> promoter generally provided a higher level of constitutive β-glucuronidase accumulation than the viral CaMV <i>35S</i> promoter. Although the <i>StLhca3</i> promoter was predominantly expressed in a green tissue-specific manner (leaves and stems) while <i>StGBSS</i> and <i>StPat</i> mainly provided tuber-specific activity, a “promoter leakage” was also found. However, the degree of unspecific activity depended on the particular transgenic line and tissue. According to fluorometric data, the functional activity of promoters in leaves could be arranged as follows: <i>StLhca3</i> > <i>StUbi</i> > CaMV <i>35S</i> > <i>StPat</i> > <i>StGBSS</i> (from highest to lowest). In tubers, the higher expression was detected in transgenic plants expressing <i>StPat-gusA</i> fusion construct, and the strength order was as follows: <i>StPat</i> > <i>StGBSS</i> > <i>StUbi</i> > CaMV <i>35S</i> > <i>StLhca3</i>. The observed differences between expression patterns are discussed considering the benefits and limitations for the usage of each promoter to regulate the expression of genes in a particular potato tissue.