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A Root Tip-Specific Expressing Anthocyanin Marker for Direct Identification of Transgenic Tissues by the Naked Eye in Symbiotic Studies.

ABSTRACT: The Agrobacterium rhizogenes hairy root transformation system is widely used in symbiotic studies of model legumes. It typically relies on fluorescent reporters, such as DsRed, for identification of transgenic roots. The MtLAP1 transcription factor has been utilized as a reporter system in Medicago truncatula based on production of anthocyanin pigment. Here, we describe a version of this reporter driven by a root-cap specific promoter for direct observation of anthocyanin accumulation in root tips, which allows the identification of transgenic hairy roots by the naked eye. Results from our analysis suggest that the reporter had no significant effects on nodulation of M. truncatula. This approach, by virtue of its strong and specific expression in root cap cells, greatly reduces false positives and false negatives, and its use of an easily scored visible pigment should allow greater versatility and efficiency in root biology studies.

SUBMITTER: Ruan Y

PROVIDER: S-EPMC8004629 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:The cultivated potato Solanum tuberosum L. is the fourth most important crop worldwide. Anthocyanins synthesis and accumulation in potato tissues are considered as one of important traits related to stress resistance and nutritional value. It is considered that the major regulatory gene for anthocyanin biosynthesis is R2R3 MYB-encoding gene StAN1. However, the genetic control of pigmentation of different potato tissues is substantially under investigated. The development of genetic markers for breeding of potato with specific pigmentation pattern remains an actual task. RESULTS:We investigated 36 potato varieties and hybrids with different pigmentation of tubers and leaves. Sequence organization of regulatory R2R3 MYB (StAN1, StMYBA1, StMYB113), bHLH (StbHLH1, StJAF13) and WD40 (StWD40) genes potentially controlling anthocyanin biosynthesis has been evaluated. The results demonstrated a high variability in the StAN1 third exon and promoter region with the exception for 35?bp, containing elements for the transcription start and activation of gene expression in roots. The analysis of transcriptional activity of genes coding R2R3 MYBs, bHLHs and WD40 transcriptional factors in leaves of eight potato genotypes with different anthocyanin pigmentation was performed. The results showed a relation between the gene expression level and plant pigmentation only for the StAN1 and StWD40 genes, while other studied genes had either strong expression in all varieties and hybrids (StMYBA1, StbHLH1 and StJAF13) or they were not expressed at all (StMYB113). CONCLUSIONS:It was found that StAN1 is the major regulatory gene controlling potato anthocyanin synthesis. However, diagnostic markers developed for the functional StAN1 alleles (StAN1777 and StAN1816) can not be used efficiently for prediction of potato pigmentation patterns. It is likely that the sequence organization of StAN1 promoter is important for anthocyanin synthesis control and the development of additional diagnostic markers is necessary.

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A Root Tip-Specific Expressing Anthocyanin Marker for Direct Identification of Transgenic Tissues by the Naked Eye in Symbiotic Studies.

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