Method for the preparation of transgenic plants characterised by geminivirus lasting resistance

A method for the preparation of transgenic plants, plant tissue or cells thereof having long lasting resistance against geminiviruses, including the following steps:

• a) selection of a geminivirus C1/AL1/AC1 polynucleotide sequence encoding an amino acid sequence conferring resistance against geminiviruses, said resistance being overcome over time by geminivirus induced post-transcriptional transgene silencing;
• b) mutagenesis of the selected geminivirus C1/AL1/AC1 polynucleotide sequence, wherein the mutations consist of silent point mutations distributed along the geminivirus C1/AL1/AC1 polynucleotide sequence so that continuous homology between the mutated C1/AL1/AC1 polynucleotide sequence and the selected C1/AL1/AC1 polynucleotide sequence is less than or equal to 8 nucleotides;
• and
• c) insertion of the geminivirus C1/AL1/AC1 polynucleotide sequence mutated in the step b) in the plant, plant tissue or cell thereof, using a construct comprising an heterologous polynucleotide sequence containing in the 5′-3′ direction:
• i) a polynucleotide sequence acting as a promoter in said plant, plant tissue or cells;
• ii) a non-translated polynucleotide sequence positioned 5′ of the encoding region of the mutated geminivirus C1/AL1/AC1 polynucleotide sequence;
• iii) a mutated C1/AL1/AC1 polynucleotide sequence mutagenized according to step (b) encoding a geminivirus amino acid sequence, wherein the mutated C1/AL1/AC1 polynucleotide sequence is an ineffective target of the geminivirus induced post-transcriptional transgene silencing;
• and
• iv) a polynucleotide sequence acting as a transcription terminator positioned 3′ with respect to said mutated C1/AL1/AC1 polynucleotide sequence.