Vertebrate intron signals comprise the 5' and 3' splice site, a branchpoint sequence and a polypyrimidine tract downstream of the branchpoint. Plant introns contain 5' and 3' splice sites similar to those of vertebrate and yeast introns. However, plant introns are different in being U-rich.

To analyse intron splicing signals, mutations are cloned into an expression cassette, transfected into protoplasts, RNA isolated and analysed by RT-PCR. By using one fluorescently labelled primer, products are separated on automatic DNA sequencing gels.

We have demonstrated that branchpoints are required for efficient intron splicing through mutational analysis. We have also used the sensitive potato invertase mini-exon system to systematically mutate the sequence surrounding the branchpoint and the polypyrimidine tract. This analysis has defined a plant branchpoint and polypyrimidine tract for the first time. The sequences (below) are similar to vertebrate sequences:

Brown (1996) Plant J. 10, 771-780.
Brown et al. (1996) Plant Mol. Biol. 32, 531-535.
Simpson et al. (1996) Plant J. 9, 369-380.
Simpson et al. (2002) RNA 8, 47-56.
Simpson et al. (2004) Plant J. 37, 82-91.