Dataset Information

Evaluation of Plant-Derived Promoters for Constitutive and Tissue-Specific Gene Expression in Potato.

ABSTRACT: 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 ?-glucuronidase (gusA) gene. The strengths of StUbi, StGBSS, StPat, and StLhca3 promoters were compared with the conventional constitutive CaMV 35S 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 StUbi promoter generally provided a higher level of constitutive ?-glucuronidase accumulation than the viral CaMV 35S promoter. Although the StLhca3 promoter was predominantly expressed in a green tissue-specific manner (leaves and stems) while StGBSS and StPat 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: StLhca3 > StUbi > CaMV 35S > StPat > StGBSS (from highest to lowest). In tubers, the higher expression was detected in transgenic plants expressing StPat-gusA fusion construct, and the strength order was as follows: StPat > StGBSS > StUbi > CaMV 35S > StLhca3. 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.

SUBMITTER: Miroshnichenko D

PROVIDER: S-EPMC7696379 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Evaluation of Plant-Derived Promoters for Constitutive and Tissue-Specific Gene Expression in Potato.

Miroshnichenko Dmitry D Firsov Aleksey A Timerbaev Vadim V Kozlov Oleg O Klementyeva Anna A Shaloiko Lyubov L Dolgov Sergey S

Plants (Basel, Switzerland) 20201109 11

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 (gusA) gene. The strengths of StUbi, StGBSS, StPat, and StLhca3 promoters were compared with the conventional constitutive CaMV 35S promoter in various organs (leaves, stems, roots ...[more]

PMID: 33182387

Similar Datasets

Project description:Streptomycetes attract a lot of attention in metabolic engineering and synthetic biology because of their well-known ability to produce secondary metabolites. However, the available constitutive promoters are rather limited in this genus.In this work, constitutive promoters were selected from a pool of promoters whose downstream genes maintained constant expression profiles in various conditions. A total of 941 qualified genes were selected based on systematic analysis of five sets of time-series transcriptome microarray data of Streptomyces coelicolor M145 cultivated under different conditions. Then, 166 putative constitutive promoters were selected by following a rational selection workflow containing disturbance analysis, function analysis, genetic loci analysis, and transcript abundance analysis. Further, eight promoters with different strengths were chosen and subjected to experimental validation by green fluorescent protein reporter and real-time reverse-transcription quantitative polymerase chain reaction in S. coelicolor, Streptomyces venezuelae and Streptomyces albus. The eight promoters drove the stable expression of downstream genes in different conditions, implying that the 166 promoters that we identified might be constitutive under the genus Streptomyces. Four promoters were used in a plug-and-play platform to control the expression of the cryptic cluster of jadomycin B in S. venezuelae ISP5230 and resulted in different levels of the production of jadomycin B that corresponded to promoter strength.This work identified and evaluated a set of constitutive promoters with different strengths in streptomycetes, and it enriched the presently available promoter toolkit in this genus. These promoters should be valuable in current platforms of metabolic engineering and synthetic biology for the activation of cryptic biosynthetic clusters and the optimization of pathways for the biosynthesis of important natural products in Streptomyces species.

Project description:Transcriptome diversity provides the key to cellular identity. One important contribution to expression diversity is the use of alternative promoters, which creates mRNA isoforms by expanding the choice of transcription initiation sites of a gene. The proximity of the basal promoter to the transcription initiation site enables prediction of a promoter's location based on the gene annotations. We show that annotation of alternative promoters regulating expression of transcripts with distinct first exons enables a novel methodology to quantify expression levels and tissue specificity of mRNA isoforms.The use of distinct alternative first exons in 3,296 genes was examined using exon-microarray data from 11 human tissues. Comparing two transcripts from each gene we found that the activity of alternative promoters (i.e., P1 and P2) was not correlated through tissue specificity or level of expression. Furthermore neither P1 nor P2 conferred any bias for tissue-specific or ubiquitous expression. Genes associated with specific diseases produced transcripts whose limited expression patterns were consistent with the tissue affected in disease. Notably, genes that were historically designated as tissue-specific or housekeeping had alternative isoforms that showed differential expression. Furthermore, only a small number of alternative promoters showed expression exclusive to a single tissue indicating that "tissue preference" provides a better description of promoter activity than tissue specificity. When compared to gene expression data in public databases, as few as 22% of the genes had detailed information for more than one isoform, whereas the remainder collapsed the expression patterns from individual transcripts into one profile.We describe a computational pipeline that uses microarray data to assess the level of expression and breadth of tissue profiles for transcripts with distinct first exons regulated by alternative promoters. We conclude that alternative promoters provide individualized regulation that is confirmed through expression levels, tissue preference and chromatin modifications. Although the selective use of alternative promoters often goes uncharacterized in gene expression analyses, transcripts produced in this manner make unique contributions to the cell that requires further exploration.

Dataset Information

Evaluation of Plant-Derived Promoters for Constitutive and Tissue-Specific Gene Expression in Potato.

Publications

Evaluation of Plant-Derived Promoters for Constitutive and Tissue-Specific Gene Expression in Potato.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets