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Marker-free PLRV resistant potato mediated by Cre-loxP excision and RNAi

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Title Marker-free PLRV resistant potato mediated by Cre-loxP excision and RNAi
 
Creator Orbegozo, Jeanette
Solorzano, Dennis
Cuéllar, Wilmer Jose
Bartolini, Maria Lupe
Ghislain, M.
Kreuze, Jan F.
 
Subject potato leaf roll luteovirus
genetic transformation
viruses
hybridization
transgenic plants
virus
transformación genética
hibridación
plantas transgénicas
 
Description An inverted repeat construct corresponding to a segment of the potato leaf roll virus coat protein gene was created under control of a constitutive promoter and transferred into a transformation vector with a heat inducible Cre-loxP system to excise the nptII antibiotic resistance marker gene. Fifty-eight transgenic events were evaluated for resistance to PLRV by greenhouse inoculations, which lead to the identification of 7 highly resistant events, of which 4 were extremely resistant. This resistance was also highly effective against accumulation in subsequent tuber generations from inoculated plants, which has not been reported before. Northern blot analysis showed correlation of PLRV specific siRNA accumulation with the level of PLRV resistance. Heat mediated excision of the nptII antibiotic resistance gene in PLRV resistant events was highly efficient in one event with full excision in 71 % of treated explants. On the other hand 8 out of 10 analyzed events showed truncated T-DNA insertions lacking one of the two loxP sites as determined by PCR and confirmed by sequencing flanking regions in 2 events, suggesting cryptic LB sites in the non-coding region between the nptII gene and the flanking loxP site. Accordingly, it is proposed to modify the Cre-loxP vector by reducing the 1 kb size of the region between nptII, loxP, and the LB.
 
Date 2016-12
2016-08-29T18:27:26Z
2016-08-29T18:27:26Z
 
Type Journal Article
 
Identifier Orbegozo, Jeanette; Solorzano, Dennis; Cuellar, Wilmer J.; Bartolini, Ida; Roman, Maria Lupe; Ghislain, Marc; Kreuze, Jan. 2016. Marker-free PLRV resistant potato mediated by Cre-loxP excision and RNAi . Transgenic Research 25(6): 813-828.
0962-8819
https://hdl.handle.net/10568/76626
https://doi.org/10.1007/s11248-016-9976-y
 
Language en
 
Rights CC-BY-4.0
Open Access
 
Format 813-828
 
Publisher Springer Science and Business Media LLC
 
Source Transgenic Research