Towards the isolation of resistance genes by transposon targeting in potato

Pereira, Andy; Jacobs, J.M.E.; Lintel-Hekkert, W. te; Rutgers, E.; Jacobsen, E.; Stiekema, Willem J.

doi:10.1007/bf01974488

Cited by 9 publications

(3 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Functional discovery of genes in potato is still a lengthy process and is often hampered by the complex characteristics associated with the potato genome, including autotetraploidy, self-incompatibility, and high heterozygosity. Although several gene discovery tools have been used in potato research, including transposon-based insertional mutagenesis [1] , [2] , gene activation-tagging [3] , and map-based cloning [4] , [5] , applications of these techniques were time-consuming, resource-intensive, and technically challenging. RNA interference (RNAi)-based potato gene silencing has recently been reported by several laboratories [6] , [7] , [8] , [9] , [10] , [11] , [12] , [13] .…”

Section: Introductionmentioning

confidence: 99%

Agrobacterium-Mediated Transient Gene Expression and Silencing: A Rapid Tool for Functional Gene Assay in Potato

et al. 2009

View full text Add to dashboard Cite

Potato is the third most important food crop worldwide. However, genetic and genomic research of potato has lagged behind other major crops due to the autopolyploidy and highly heterozygous nature associated with the potato genome. Reliable and technically undemanding techniques are not available for functional gene assays in potato. Here we report the development of a transient gene expression and silencing system in potato. Gene expression or RNAi-based gene silencing constructs were delivered into potato leaf cells using Agrobacterium-mediated infiltration. Agroinfiltration of various gene constructs consistently resulted in potato cell transformation and spread of the transgenic cells around infiltration zones. The efficiency of agroinfiltration was affected by potato genotypes, concentration of Agrobacterium, and plant growth conditions. We demonstrated that the agroinfiltration-based transient gene expression can be used to detect potato proteins in sub-cellular compartments in living cells. We established a double agroinfiltration procedure that allows to test whether a specific gene is associated with potato late blight resistance pathway mediated by the resistance gene RB. This procedure provides a powerful approach for high throughput functional assay for a large number of candidate genes in potato late blight resistance.

show abstract

Section: Introductionmentioning

confidence: 99%

Agrobacterium-Mediated Transient Gene Expression and Silencing: A Rapid Tool for Functional Gene Assay in Potato

et al. 2009

View full text Add to dashboard Cite

show abstract

“…Although T‐DNA insertional mutagenesis and transposon tagging have been reported in dihaploid potato (EIKharbotly et al ., 1996; Pereira et al ., 1992; Van Enckevort et al ., 2000, 2001), this approach is time consuming because interesting material (usually tetraploid) has to be taken first to the diploid level. When tetraploid Solanum tuberosum is diploidised, through either anther culture or the use of certain dihaploid inducer S. phureja clones (Ortiz and Peloquin, 1994), a powerful gametophytic self incompatibility system (Clarke et al ., 1989; Hermsen, 1978; Pushkarnath, 1942) complicates further genetic analyses.…”

Section: Introductionmentioning

confidence: 99%

Virus‐induced gene silencing in Solanum species

et al. 2004

View full text Add to dashboard Cite

Both authors contributed equally to this work. SummaryVirus-induced gene silencing (VIGS) has been used routinely in Nicotiana benthamiana to assess functions of candidate genes and as a way to discover new genes required for diverse pathways, especially disease resistance signalling. VIGS has recently been shown to work in Arabidopsis thaliana and in tomato. Here, we report that VIGS using the tobacco rattle virus (TRV) viral vector can be used in several Solanum species, although the choice of vector and experimental conditions vary depending on the species under study. We have successfully silenced the phytoene desaturase (PDS) gene in the diploid wild species Solanum bulbocastanum and S. okadae, in the cultivated tetraploid S. tuberosum and in the distant hexaploid relative S. nigrum (commonly known as deadly nightshade). To test whether the system could be utilised as a rapid way to assess gene function of candidate resistance (R) genes in potato and its wild relatives, we silenced R1 and Rx in S. tuberosum and RB in S. bulbocastanum. Silencing of R1, Rx and RB successfully attenuated R-gene-mediated disease resistance and resulted in susceptible phenotypes in detached leaf assays. Thus, the VIGS system is an effective method of rapidly assessing gene function in potato.

show abstract

“…By using a combination of approaches, i.e., chromosome tagging with selectable markers, such as an^ or H~~ through transposon tagging, as has been realised for the whole haploid set of potato chromosomes by Pereira et al (1992); or by one to one microfusion (Spangenberg et al 199 l ) , which needs no selectable markers; the MMCT technique can facilitate the construction of a complete set of monosomic addition plants. The direct production of monosomic additions in sexually incongruent plant species is potentially useful for the transfer of economically important traits, such as disease resistances that are controlled by polygenes clustered within blocks on specific chromosomes Genome Downloaded from www.nrcresearchpress.com by Laurentian University on 11/05/14…”

Section: Discussionmentioning

confidence: 99%

Microprotoplast-mediated transfer of single specific chromosomes between sexually incompatible plants

Ramulu¹,

Dijkhuis²,

Rutgers³

et al. 1996

Genome

Self Cite

View full text Add to dashboard Cite

Microprotoplast-mediated chromosome transfer (MMCT) through fusion of small (subdiploid) microprotoplasts of a transgenic triploid potato (Solanum tuberosum) cell line with leaf protoplasts of tobacco (Nicotiana tabacum) and the wild tomato species Lycopersicon peruvianum is reported. The microprotoplasts contained one or a few chromosomes. Monosomic addition plants were produced from the fusion products. We employed mass-scale induction of micronuclei in donor suspension cells of potato using the microtubule inhibitor Cremart. Protoplasts were isolated from micronucleated cells after incubation in a cell wall digesting enzyme mixture. The microprotoplasts were isolated from the micronucleated protoplasts by high-speed centrifugation. By using sequential filtration, small microprotoplasts containing one or few chromosomes were separated from the bigger subdiploid microprotoplasts. These small microprotoplasts were fused with recipient protoplasts of tobacco or tomato using polyethylene glycol. The selectable marker kanamycin resistance (KanR) and the reporter gene β-glucuronidase (gus), carried by the donor potato chromosome, were used for the selection of fusion products and the isolation of hybrid calli. Several monosomic addition plants were obtained within the short period of 3-4 months after fusion. These contained one potato chromosome carrying a single copy of gus and one or two copies of the neomycin phosphotransferase (nptII) gene conferring KanR, and the complete set of chromosomes of tobacco or tomato, as revealed by genomic in situ hybridization and Southern blot hybridization. The alien genes, gus and nptII, were stably expressed in both the tobacco and tomato backgrounds. They were transmitted to the progeny after backcrossing to tomato. Monosomic and disomic additions, and some introgression plants showing integration of gus and nptII in the tomato genome, were recovered in the first backcross progeny. The potential value of MMCT for the transfer of economically important traits, genome analysis, and gene expression is discussed. Key words : chromosome transfer, microprotoplast fusion, monosomic-disomic additions, sexual transmission, DNA integration, alien gene expression.

show abstract

Towards the isolation of resistance genes by transposon targeting in potato

Cited by 9 publications

References 28 publications

Agrobacterium-Mediated Transient Gene Expression and Silencing: A Rapid Tool for Functional Gene Assay in Potato

Agrobacterium-Mediated Transient Gene Expression and Silencing: A Rapid Tool for Functional Gene Assay in Potato

Virus‐induced gene silencing in Solanum species

Microprotoplast-mediated transfer of single specific chromosomes between sexually incompatible plants

Contact Info

Product

Resources

About