A rapid and efficient Agrobacterium-mediated transient gene expression system for strawberry leaves and the study of disease resistance proteins

Cui, Meng-Yuan; Wei, Wei; Gao, Kuan; Xie, Yeming; Guo, Yifeng; Feng, Jiahao

doi:10.1007/s11240-017-1279-3

Cited by 15 publications

(7 citation statements)

References 43 publications

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“…Two NBS-RPW8 proteins conferring mildew-resistance have been described in the Arabidopsis thaliana genome (Xiao et al 2001) that retained their function when expressed in grape (Hu et al 2018). The At RPW8.2 gene was recently shown to induce the expression of defense-related genes when expressed in strawberry leaves (Cui et al 2017). This R-gene subtype has apparently expanded in strawberry, possibly due to unusually high mildew disease pressure exerted on strawberry species and intense selection for resistance.…”

Section: Discussionmentioning

confidence: 99%

Disease Resistance Genetics and Genomics in Octoploid Strawberry

Barbey

Lee

Verma

et al. 2019

G3 Genes|Genomes|Genetics

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Octoploid strawberry (Fragaria ×ananassa) is a valuable specialty crop, but profitable production and availability are threatened by many pathogens. Efforts to identify and introgress useful disease resistance genes (R-genes) in breeding programs are complicated by strawberry’s complex octoploid genome. Recently-developed resources in strawberry, including a complete octoploid reference genome and high-resolution octoploid genotyping, enable new analyses in strawberry disease resistance genetics. This study characterizes the complete R-gene collection in the genomes of commercial octoploid strawberry and two diploid ancestral relatives, and introduces several new technological and data resources for strawberry disease resistance research. These include octoploid R-gene transcription profiling, dN/dS analysis, expression quantitative trait loci (eQTL) analysis and RenSeq analysis in cultivars. Octoploid fruit eQTL were identified for 76 putative R-genes. R-genes from the ancestral diploids Fragaria vesca and Fragaria iinumae were compared, revealing differential inheritance and retention of various octoploid R-gene subtypes. The mode and magnitude of natural selection of individual F. ×ananassa R-genes was also determined via dN/dS analysis. R-gene sequencing using enriched libraries (RenSeq) has been used recently for R-gene discovery in many crops, however this technique somewhat relies upon a priori knowledge of desired sequences. An octoploid strawberry capture-probe panel, derived from the results of this study, is validated in a RenSeq experiment and is presented for community use. These results give unprecedented insight into crop disease resistance genetics, and represent an advance toward exploiting variation for strawberry cultivar improvement.

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Section: Discussionmentioning

confidence: 99%

Disease Resistance Genetics and Genomics in Octoploid Strawberry

Barbey

Lee

Verma

et al. 2019

G3 Genes|Genomes|Genetics

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“…Similarly, leaf age affected transformation efficiency in cacao (Theobroma cacao L) (Fister et al 2016) and strawberry (Fragaria vesca) (Cui et al 2017). Transient expression of yfp and gfp genes decreased with increasing leaf age (Fister et al 2016;Cui et al 2017). Fister et al (2016) attribute this result to both physiological and physical (toughness, rigidity) differences between leaves of different ages.…”

Section: Effect Of Plant Age On Transient Gus Expressionmentioning

confidence: 99%

A simple and efficient agroinfiltration method in coffee leaves (Coffea arabica L.): assessment of factors affecting transgene expression

et al. 2018

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The establishment of a simple, rapid and efficient transient expression system is a necessary tool for the functional validation of candidate genes in coffee biotechnology. The effects of Agrobacterium strain, age of the donor plant, infiltration method, and infiltration medium on transgene expression in detached coffee leaves were evaluated. Regarding the effect of Agrobacterium strain, the expression of uidA was higher in GV3101-treated coffee disks than in LBA4404 and ATHV-treated samples. On the other hand, transient expression of uidA was significantly higher in leaf disks from young plants (6-weeks-old) (13.1 ± 1.4%) than in mature tissue (12-weeks-old) (1.6 ± 1.2%). Transient uidA expression was higher in detached coffee leaf disks from young plants infiltrated with one injection of 15 µL of Agrobacterium strain GV3101::1303 suspended in MS salts supplemented with 30 g/L sucrose, 1.9 g/L MES and 200 uM AS with subsequent sanding of the abaxial epidermis. Using the optimized protocol, expression of the uidA gene was observed 6, 24 and 48 h and 5 weeks after bacterial injection. DNA was extracted from coffee disks with positive GUS expression and specific mgfp5 and uidA fragments were amplified 5 weeks post-agroinfiltration. On the other hand, using the optimized protocol, a specific cry10Aa (500 bp) fragment was amplified in the agro-infiltrated coffee leaf disks 5 weeks post-agroinfiltration with the plasmid pB427-35S-cry10Aa. Moreover, the expression of the gene cry10Aa in two infiltrated coffee leaf disks was verified by RT-PCR and an expected 500 bp fragment was amplified.

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“…Agrobacterium-mediated transient transformation was used in many angiosperm plants to get transient and high-level expression of target genes, such as in Arabidopsis, rice, wheat, Nicotiana benthamiana, strawberry, and soybean [7,[34][35][36][37]. The first successful Agrobacterium-mediated transformation of conifer was in Sugar pine has the longest cones of any conifer.…”

Section: Discussionmentioning

confidence: 99%

An efficient system for Agrobacterium-mediated transient transformation in Pinus tabuliformis

Liu

et al. 2020

Plant Methods

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Background: Functional genomic studies using genetics approaches of conifers are hampered by the complex and enormous genome, long vegetative growth period, and exertion in genetic transformation. Thus, the research carried out on gene function in Pinus tabuliformis is typically performed by heterologous expression based on the model plant Arabidopsis. However, due to the evolutionary and vast diversification from non-flowering (gymnosperms) to flowering (angiosperms) plants, several key differences may alter the underlying genetic concerns and the analysis of variants. Therefore, it is essential to develop an efficient genetic transformation and gene function identification protocol for P. tabuliformis. Results: In the present study we established a highly efficient transgene Agrobacterium-mediated transient expression system for P. tabuliformis. Using a β-glucuronidase gene (GUS) as a reporter gene expression, the highest transformation efficiency (70.1%) was obtained by co-cultivation with Agrobacterium strain GV3101 at an optical density at 600 nm of 0.8, with 150 μM acetosyringone for 30 min followed by 3 days in the dark at 23 ± 1 °C. This protocol would be applied to other conifers; GUS staining was observed 24 h post-infection. Conclusions: We report a simple, fast, and resilient system for transient Agrobacterium-mediated transformation high-level expression of target genes in P. tabuliformis, which will also improve transformation efficiency in other conifer species.

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A rapid and efficient Agrobacterium-mediated transient gene expression system for strawberry leaves and the study of disease resistance proteins

Cited by 15 publications

References 43 publications

Disease Resistance Genetics and Genomics in Octoploid Strawberry

Disease Resistance Genetics and Genomics in Octoploid Strawberry

A simple and efficient agroinfiltration method in coffee leaves (Coffea arabica L.): assessment of factors affecting transgene expression

An efficient system for Agrobacterium-mediated transient transformation in Pinus tabuliformis

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