PCR-based cloning and segregation analysis of functional gene homologues in Beta vulgaris

Abstract

To analyse genetic factors that potentially affect sugar quality and yield in Beta vulgaris, we designed primers based on 18 homologous ESTs and conserved regions of 32 heterologous ESTs encoding gene products that act in the Calvin cycle, the oxidative pentose phosphate cycle, photorespiration, synthesis, transport and degradation of sucrose, glycolysis, the citric acid cycle, nitrogen metabolism and osmoprotection. Data on the amplification of 54 gene homologues from B. vulgaris are presented. Among these are 35 homologues for which DNA sequence information from B. vulgaris is now available for the first time. For genetic mapping a PCR-based strategy using CAPS (cleaved amplified polymorphic sequence), DFLP (DNA fragment length polymorphism), SSCP (single-strand conformation polymorphism) and HD (heteroduplex) analysis was adopted. RFLP analysis was also used in some cases. The different techniques used for the detection of polymorphisms are evaluated with respect to their sensitivity and versatility. In all, 42 functional genes have been assigned to the nine linkage groups of sugar beet.