Conversion of AFLP markers to sequence-specific PCR markers in barley and wheat

Conversion of amplified fragment length polymorphisms (AFLPs) to sequence-s pecific PCR primers would be useful for many genetic-linkage applications. We examined 21 wheat nullitetrasomic stocks and five wheat-barley addition lines using 12 and 14 AFLP primer combinations, respectively. On average, 3 6.8% of the scored AFLP fragments in the wheat nullitetrasomic stocks and 2 2.3% in the wheat-barley addition lines could be mapped to specific chromos omes, providing approximately 461 chromosome-specific AFLP markers in the w heat nullitetrasomic stocks and 174 in the wheat-barley addition lines. Ten AFLP fragments specific to barley chromosomes and 16 AFLP fragments specif ic to wheat 3BS and 4BS chromosome arms were isolated from the polyacrylami de gels, re-amplified, cloned and sequenced. Primer sets were designed from these sequences. Amplification of wheat and barley genomic DNA using the b arley derived primers revealed that three primer sets amplified DNA from th e expected chromosome, five amplified fragments from all barley chromosomes but not from wheat, one amplified a similar-sized fragment from multiple b arley chromosomes and from wheat, and one gave no amplification. Amplificat ion of wheat genomic DNA using the wheat-derived primer sets revealed that three primer sets amplified a fragment from the expected chromosome, 11 pri mer sets amplified a similar-sized fragment from multiple chromosomes, and two gave no amplification. These experiments indicate that polymorphisms id entified by AFLP are often not transferable to more sequence-specific PCR a pplications.