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Anthocyanin production as a potential visual selection marker during plant transformation.

ABSTRACT: A mutant allele of the transcription factor gene MYB10 from apple induces anthocyanin production throughout the plant. This gene, including its upstream promoter, gene coding region and terminator sequence, was introduced into apple, strawberry and potato plants to determine whether it could be used as a visible selectable marker for plant transformation as an alternative to chemically selectable markers, such as kanamycin resistance. After transformation, red coloured calli, red shoots and red well-growing plants were scored. Red and green shoots were harvested from apple explants and examined for the presence of the MYB10 gene by PCR analysis. Red shoots of apple explants always contained the MYB10 gene but not all MYB10 containing shoots were red. Strawberry plants transformed with the MYB10 gene showed anthocyanin accumulation in leaves and roots. No visible accumulation of anthocyanin could be observed in potato plants grown in vitro, even the ones carrying the MYB10 gene. However, acid methanol extracts of potato shoots or roots carrying the MYB10 gene contained up to four times higher anthocyanin content than control plants. Therefore anthocyanin production as result of the apple MYB10 gene can be used as a selectable marker for apple, strawberry and potato transformation, replacing kanamycin resistance.

SUBMITTER: Kortstee AJ

PROVIDER: S-EPMC3210953 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundArsenic is a carcinogen that targets the urogenital system, including the prostate. Although the mechanisms for arsenic-induced carcinogenesis are undefined, arsenic drives overaccumulation of stem cells and cancer stem cells (CSCs) in vivo and in vitro, indicating that these cells are a key target population. Disruption of stem cell population dynamics may be critical to acquisition of cancer phenotype. We tested the hypothesis that prostate stem cells have a survival selection advantage during arsenic exposure that favors their accumulation and facilitates their malignant transformation.MethodsInnate and acquired resistance to acute (24-72 hours of exposure) and chronic (6 weeks of exposure) arsenite-induced cytolethality and apoptosis were assessed in a human prostate stem cell line (WPE-stem) and the mature parental cell line (RWPE-1). Real-time reverse transcription-polymerase chain reaction and/or Western blot analysis was used to measure the expression of apoptosis-, stress-, and arsenic-related genes. Arsenic-, cadmium-, and N-methyl-N-nitrosourea-induced isogenic malignant transformants of RWPE-1 cells were compared for acquisition of CSC-like qualities by holoclone and sphere formation assays, growth in soft agar, and expression of CSC biomarkers. All statistical tests were two-sided.ResultsWPE-stem cells showed innate resistance to arsenic-induced cytolethality (arsenite concentration lethal to 50% of the cells [LC(50)] = 32.4 microM, 95% confidence interval [CI] = 31.5 to 33.3 muM) and apoptosis compared with parental RWPE-1 cells (LC(50) = 10.4 muM, 95% CI = 7.4 to 13.4 microM). Compared with RWPE-1 cells, WPE-stem cells showed noticeably higher expression of antiapoptotic (ie, BCL2, MT), stress-related (ie, NFE2L2, SOD1, PRODH), and arsenic adaptation (ie, ABCC1, GSTP1) factors and noticeably lower expression of proapoptotic factors (ie, BAX, caspases 3, 7, 8, and 9). WPE-stem cells also showed hyper-adaptability to chronic arsenite exposure (5 microM, 6 weeks) compared with RWPE-1 cells (LC(50) = 94.7 vs 32.1 microM, difference = 62.6 muM, 95% CI = 53.3 to 71.9 muM) at levels that in previous work induced a malignant phenotype in RWPE-1 after 30 weeks of exposure. Quantification of CSC-like cells in isogenic RWPE-1 transformants showed that marked overproduction was unique to a malignant phenotype acquired in response to arsenic exposure but not in response to cadmium or N-methyl-N-nitrosourea exposure.ConclusionsAn apparent stem cell survival advantage with regard to arsenic causes selection during malignant transformation that manifests itself as an overabundance of CSC-like cells specifically after arsenic-driven acquisition of malignant phenotype. The increased resistance to apoptosis and arsenite hyper-adaptability of WPE-stem cells suggests that arsenite transformation of RWPE-1 cells involves an increase in the number of CSC-like cells.

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Anthocyanin production as a potential visual selection marker during plant transformation.

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