Date of Award

Winter 2001

Project Type

Dissertation

Program or Major

Plant Biology

Degree Name

Doctor of Philosophy

First Advisor

Thomas M Davis

Abstract

A candidate gene approach was used to determine the likely molecular identity of the c locus (yellow fruit color) in Fragaria vesca, a diploid (2n = 2x = 14) strawberry. Using PCR with degenerate primer pairs, intron-containing segments of structural genes coding for the enzymes: chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanindin synthase (ANS), and one Del-like regulatory gene (RAN) in the anthocyanin biosynthetic pathway were amplified, cloned and sequenced. Intron length polymorphisms for each of these genes were detected among three diploid varieties: F. vesca Alpine variety 'Yellow Wonder' (YW) (Europe); DNIC, an F. vesca clone collected from Northern California; and F. nubicola FRA520, a U.S.D.A. accession collected in Pakistan. Using F2 generations of the crosses DNIC x YW and YW x FRA520 as mapping populations, the six candidate genes were mapped in relation to previously mapped randomly amplified polymorphic DNA (RAPD) markers and morphological markers. The F3H gene was linked without recombination to the c locus in linkage group I, while the other five candidate genes mapped to different linkage groups. These results suggest that the wild type allele (C) of the c (yellow fruit color) locus encodes an F3H necessary for red fruit in F. vesca.

Further study and characterization of the F3H gene in strawberry was carried out. Using 5'-RACE and 3 '-RACE technologies, two overlapping F3H cDNAs which representing the complete F3H cDNA sequence were isolated and cloned from the fruit of commercial strawberry, Fragaria x ananassa CC2, an octoploid species provided by U.S.D.A. Based on the F3H cDNA sequence, new strawberry specific primers were designed to clone and sequence the full-length F3H gene in YW and DN1C genomes. Additionally, promoter sequence of the F3H gene was also cloned and sequenced from YW, DN1C and FRA364, using a modified inverse PCR technique. The alignments of these sequences found two possible mutation sites, one a base substitution resulting in a K → T amino acid substitution, and the other in the promoter region. These results are consistent with identity between the c locus and the F3H gene, but further studies are needed to confirm this hypothesis.

An intron polymorphism in intron I of the F3H gene was detected between YW and other red-fruited species, and was used to map the F3H gene in new populations: F1 population of cross YW x F1 (YW x FRA520) and F2 population of cross YW x FRA364. Based on F2 population of the interspecific cross YW x FRA364, a red versus white internal fruit color polymorphism also mapped to the locus of the F3H gene, as well as c locus. This result further implicates the F3H gene as an important contributor to fruit color variation in strawberry.

Download

Share

COinS