Molecular cloning of the self-incompatibility genes S1 and S3 from almond ( Prunus dulcis cv. Ferragn�s)

Abstract

Almond (Prunus dulcis) displays gametophytic self-incompatibility. In the work reported here, we cloned two novel S-RNase genes from almond cultivar Ferragnès (genotype S1S3) using PCR. The S1-RNase gene has the same coding region as the Sb gene cloned from almond cultivated in the USA; however, their introns are different in sequence. S1 was cloned and sequenced from six different cultivars originating in Europe. The full-length of the S3-RNase gene was cloned using two primers corresponding to the start and stop codons contexts. Two introns are present in the S3 gene, unique among the S-RNase genes. Sequence-specific PCR was performed to confirm that the two cloned genes co-segregate with the S-locus using progenies of a controlled cross between Tuono (S1Sf) and Ferragnès (S1S3). Based on the structural differences of S-and S-like RNase genes, we discuss the evolutionary relationship between the two groups of RNase genes.

Allele-specific PCR detection of sweet cherry self-incompatibility (S) alleles S1 to S16 using consensus and allele-specific primers

“…A rare example of intra-allelic variation in intron length that has recently been found in almond (Channuntapipat et al 2001;Ma and Oliveira 2001) indicates the need for caution in characterizing potential new alleles on the basis of intron length alone, without confirmation by sequencing and/or test crossing.…”

“…The exon sequences we obtained were not always sufficiently variable for the design of good specific primers, and the use of intron sequences was necessary in some cases. Allele-specific primers have been used in cherry ), apple (Broothaert et al 1995Janssens et al 1995Janssens et al , 1996Verdoodt et al 1998;Matsumoto and Kitahara 2000;Sakurai et al 2000;Van Nerum et al 2001) and almond (Tamura et al 2000;Channuntapipat et al 2001;Ma and Oliveira 2001), sometimes in combination with restriction enzyme digestions of PCR products, to confirm the genotype score or to distinguish between two alleles amplified with the same 'specific' primer pair. The report of intra-allelic point mutations in the coding sequence (not leading to amino-acid changes) in apple (Van Nerum et al 2001) suggests that confirmation by allele-specific restriction enzyme digestions or primers may not always be 100% reliable.…”

An apricot (Prunus armeniaca L.) F2 progeny linkage map based on SSR and AFLP markers, mapping plum pox virus resistance and self-incompatibility traits

“…Molecular typing of S-alleles has been achieved in cherry (Tao et al 1999;Sonneveld et al 2001;Wiersma et al 2001) and almond (Tao et al 1997;Ushijima et al 1998;Ma and Oliveira 2001) through the identification and characterization of cDNAs corresponding to stylar RNases. In apricot S-alleles have not been yet identified and stylar ribonuclease assays have been used to detect self-compatible seedlings in apricot (Burgos et al 1998) and other fruit tree species (Sassa et al 1992;Boskovic and Tobutt 1996;Boskovic et al 1997;Boskovic and Tobutt 1999).…”

Almond

“…Some methods of early screening have been applied to advance the selection process as well . At present, detection of self-compatibility at the seedling stage by specifi c primers (Ma and Oliveira 2001 ) allows a discarding of self-incompatible seedlings during the fi rst year of growth. Other new technologies are also being increasingly integrated in the breeding programs as summarized by Martínez-Gómez et al ( 2003b, 2005 .…”