title: Development of late blight resistant potatoes by cisgene stacking.
abstract: BACKGROUND
  Phytophthora infestans, causing late blight in potato, remains one of the most devastating pathogens in potato production and late blight resistance is a top priority in potato breeding. The introduction of multiple resistance (R) genes with different spectra from crossable species into potato varieties is required. Cisgenesis is a promising approach that introduces native genes from the crops own gene pool using GM technology, thereby retaining favourable characteristics of established varieties.
  
  
  RESULTS
  We pursued a cisgenesis approach to introduce two broad spectrum potato late blight R genes, Rpi-sto1 and Rpi-vnt1.1 from the crossable species Solanum stoloniferum and Solanum venturii, respectively, into three different potato varieties. First, single R gene-containing transgenic plants were produced for all varieties to be used as references for the resistance levels and spectra to be expected in the respective genetic backgrounds. Next, a construct containing both cisgenic late blight R genes (Rpi-vnt1.1 and Rpi-sto1), but lacking the bacterial kanamycin resistance selection marker (NPTII) was transformed to the three selected potato varieties using Agrobacterium-mediated transformation. Gene transfer events were selected by PCR among regenerated shoots. Through further analyses involving morphological evaluations in the greenhouse, responsiveness to Avr genes and late blight resistance in detached leaf assays, the selection was narrowed down to eight independent events. These cisgenic events were selected because they showed broad spectrum late blight resistance due to the activity of both introduced R genes. The marker-free transformation was compared to kanamycin resistance assisted transformation in terms of T-DNA and vector backbone integration frequency. Also, differences in regeneration time and genotype dependency were evaluated.
  
  
  CONCLUSIONS
  We developed a marker-free transformation pipeline to select potato plants functionally expressing a stack of late blight R genes. Marker-free transformation is less genotype dependent and less prone to vector backbone integration as compared to marker-assisted transformation. Thereby, this study provides an important tool for the successful deployment of R genes in agriculture and contributes to the production of potentially durable late blight resistant potatoes.
authors.forename: Kwang-Ryong
authors.initials: KR
authors.lastname: Jo
authors.forename: Chol-Jun
authors.initials: CJ
authors.lastname: Kim
authors.forename: Sung-Jin
authors.initials: SJ
authors.lastname: Kim
authors.forename: Tok-Yong
authors.initials: TY
authors.lastname: Kim
authors.forename: Marjan
authors.initials: M
authors.lastname: Bergervoet
authors.forename: Maarten A
authors.initials: MA
authors.lastname: Jongsma
authors.forename: Richard G F
authors.initials: RG
authors.lastname: Visser
authors.forename: Evert
authors.initials: E
authors.lastname: Jacobsen
authors.affiliation: Wageningen UR Plant Breeding, Wageningen University & Research Centre, P,O, Box 386, 6700 AJ Wageningen, The Netherlands. jack.vossen@wur.nl.
authors.forename: Jack H
authors.initials: JH
authors.lastname: Vossen
mesh_terms: D060054:Agrobacterium / Q000235:genetics; D060467:Disease Resistance / Q000235:genetics; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000378:metabolism; D005838:Genotype; D010641:Phenotype; D055750:Phytophthora infestans / Q000502:physiology; D010935:Plant Diseases / Q000235:genetics* / Q000382:microbiology; D010940:Plant Proteins / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics; D011198:Solanum tuberosum / Q000235:genetics*
chemical_list: D010940:Plant Proteins