Cloning And Analysis Of Fusarium Wilt Resistance Gene Homologues In Watermelon

In our study, degenerate primers including 22 upper primers and 17 lower primerswere designed based on the cloned resistance genes I₂C in tomato, Fom-2 in melon andsome RGAs in cucumber using RGA approach. 165 primer pairs were organized toscreen the RGAs relative to Fusarium wilt resistance gene in watermelon. 86 primerpairs (52%) showed polymorphic between PI296341 and 97103. A 700bp polymorphicband amplified by primer pair SF2/SR10 and a 500bp band amplified by primer pairSF12/SR5 are link to watermelon Fom-1 gene. These markers are mapped to the twoflanks of Fom-1 and the genetic distances are 25 cM and 27 cM, respectively. Primerpairs SF17/SR14, SF17/SR5, SF18/SR14 and SF18/SR5 are not polymorphic betweenthe two parents. A single band was harvested using these primer pairs and the size ofthe band is identical with the product of simulation PCR in melon Fom-2 using theseprimer pairs. 20 bands were cloned and sequenced successfully. These sequences willbe submitted to Genebank after the further confirmation. These sequences are clusteredto three groups. These deduced amino acid sequences contains the conserved motifssuch as P-loop Kinase2 and Kinase3a of NBS-kind R gene and are homologic with thecloned R gene in other plants. Seven of these sequences have intact ORFs without stopcodons. The deduced amino acids of RGA sequences 1714R3 and 175R1 harvestedfrom PI296341 containing Fom-1 and Fom-2 are homologic (62.3% and 72.5%) withFom-2 in melon. 175R1 contains a intact ORF including conserved motifs P-loop,Kinase2 and Kinase3a without stop codons. The two sequences will be of great valueto mapping and cloning Fusarium wilt resistance gene in watermelon. Specific primerscan be designed based on the difference bases between RGAs from resistant parent andthat from susceptible parent to screen for the molecular markers tightly linked toresistance gene. The breeding process will be accelerated with the benefit of MAS.