The Flavr Savr Tomato

Flavr Savr Tomato The first FDA approved genetically modified food Licensed in 1994 Ripening causes production of an enzyme Polygalacturonase in a gradual increasing level, which is responsible for softening of the tomato and which becomes the cause of rottening Calgene introduced a gene in plant which synthesize a complementary mRNA to PG gene and inhibiting the synthesis of PG enzyme.

Antisense RNA Antisense RNA (asRNA) is a single-stranded RNA that is complementary to a messenger RNA (mRNA) strand transcribed within a cell.

Put PG gene sequences in the reverse orientation into the vector Transformation Isolate the nuclei used for in vitro transcription Isolate the RNA Hybridize RNA Measure PG activity Lycopene determination

Result

PG gene suppression is the result of RNAi

Promoter antisense cDNA poly A addition signal antisense oligonucleotide mRNA -antisense RNA complex dsRNA Binding of dsRNA-specific nuclease cleavage mRNA is cleaved! Nuclease-ssRNA complex Hybridizes to mRNA sense antisense

Southern blot Two linked T-DNAs Left T-DNA, lanes 1 and 2 (primers 1 and 4); Right T-DNA, lanes 3 and 4 (primers 2 and 3); full insert, lanes 5 and 6 (primers 1 and 2); unique junction region, lanes 7 and 8 (primers 5 and 6). Diagram of the sequenced 13,621-bp T-DNA insertion and flanking genomic regions. DNA probes (bars C and D) and primers (arrows 1–6) used for confirmation PCRs are indicated.

Northern blot

Result Flavr Savr locus contains two linked T-DNAs siRNAs from transgenes accumulate in Flavr Savr tomatoes.