Hybrid-onion (Allium cepa) seed is produced using systems of cytoplasmic male sterility (CMS) and two different CMS systems have been genetically characterized. S cytoplasm was the first source of onion CMS identified in the 1920s, followed by T cytoplasm that was described in the 1960s. Numerous studies have documented polymorphisms in the organellar DNAs differentiating S and T cytoplasms from the normal male-fertile cytoplasm of onion. There may be additional source(s) of onion CMS that have been described as “T-like” and appear to be more similar to N and T cytoplasms than S cytoplasm. In this study, onion breeding lines from commercial entities were evaluated for molecular markers distinguishing sources of onion CMS. Our results reveal that bona fide T cytoplasm is rarely used commercially to produce hybrid-onion seed, and both S cytoplasm and “T-like” cytoplasm are widely used. We propose that this “T-like” cytoplasm be labeled as “R” cytoplasm because it may have originated from population(s) of ‘Rijnsburger’ onion in the Netherlands. The results of this study also help to clarify inconsistent reports regarding nuclear male-fertility restoration for different sources of onion CMS.
Michael J. Havey and Sunggil Kim
Haejeen Bang, Sunggil Kim, Daniel I. Leskovar, and Stephen King
Fruit color and carotenoid composition are important traits in watermelon. Watermelon fruit color inheritance has revealed that several genes are involved in color determination. Carotenoids are known to have various functions in plants and animals, such as providing antioxidant activity and other health benefits for humans, and UV protection and pigmentation for plants. Differential gene activity in the carotenoid biosynthetic pathway may result in different color determination of mature fruit. Eight genes encoding enzymes involved in the pathway were isolated and their structures were characterized. While obtaining full-length cDNA of these enzymes, two single-nucleotide polymorphisms were detected in a coding region of lycopene β-cyclase (LCYB). These SNP markers showed cosegregation with red and canary yellow fruit color based on the genotyping of two segregating populations. This will lead to development of a codominant molecular marker for the selection of LCYB allele, which may allow breeders to distinguish between red and canary yellow watermelon fruit colors at the seedling stage.
Ryan L. Walker, Sunggil Kim, Javier F. Betran, Kilsun Yoo, and Leonard M. Pike
Onions suffer from severe inbreeding depression, which has inhibited the development of homozygous inbred lines in breeding programs. The creation of doubled haploid (DH) lines in onion provides a unique opportunity to evaluate the utility of such lines as parents in a breeding program. For this purpose, two diallele cross experiments were conducted. The first consisted of a six-parent diallele cross using six DH lines developed at Texas A&M University. The second, a four-parent diallele cross performed with two DH lines and two inbred lines from the breeding program. Bulbs from the various crosses were evaluated for diameter, height, centers/bulb, ring thickness, number of rings/bulb, bulb weight, soluble solids content, and pungency. For some traits, general combining ability (GCA) effects explained most of the variation. However, for other traits, specific combining ability (SCA) effects predominated. For all traits, GCA and SCA were always larger than the reciprocal effects (divided into maternal and nonmaternal components). The GCA and SCA effects show an inverse correlation between the number of centers/bulb and ring thickness.
Haejeen Bang, Sunggil Kim, Daniel I. Leskovar, Angela Davis, and Stephen R. King
Gene identification and characterization can be utilized for the identification of respective functions and their relationship to flesh color inheritance. Phytoene synthase (PSY), which converts two molecules of GGPP into phytoene, is the first committed step of the pathway. Previous phylogenetic analysis of PSY has indicated that PSY duplication is common in Poaceae, but rare in dicots. Degenerate PCR and RACE were used for PSY cloning. Three members of PSY gene family (PSY-A, PSY-B and PSY-C) were identified. PSY-A shared higher identity with PSY-C than PSY-B. PSYC shared 96% identity with melon PSY. PSY-C also showed a high homology with tomato PSY1, even higher than PSY-A and PSY-B. It showed a similar gene expression pattern, so we propose that PSY-C is a homologue to PSY1. RT-PCR analysis indicated that PSY-B has a different transcriptional behavior from PSY-A, similar to tomato PSY2. Therefore, PSY genes appear to be under different regulatory mechanisms. Deduced protein sequence of PSY1 or PSY2 between species has higher homology than between PSY1 and PSY2 within species. Phylogenetic analysis indicated that watermelon PSY gene family is very distantly related. Watermelon and carrot PSY gene families did not appear to cluster as closely as in Poaceae or tomato. This indicates that watermelon and carrot PSY genes are not conserved as much as PSY in tomato or Poaceae. There was no particular pattern in phylogenetic relationship of dicots. Poaceae PSY genes showed a clustering into a PSY1 group and PSY2 group. PSY duplication in watermelon provides additional evidence that PSY duplication may be a common phenomenon in dicots. They are likely to be duplicated evolutionarily a long time ago, possibly even prior to the evolution of monocot and dicot divergence.
Sunggil Kim*, Marla Binzel, Sunghun Park, Kil-Sun Yoo, and Leonard Pike
Anthocyanin, one of the flavonoids, is a primary determinant of red color in onions. Inheritance studies indicate that a single gene determines the color difference between yellow and red onions. In order to establish which gene might be responsible for this color difference, full-length cDNAs of five structural genes: chalcone synthase (CHS), flavanone 3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), and flavonol synthase (FLS) were cloned using degenerate PCR and RACE (Rapid Amplification of cDNA Ends). RT-PCR was carried out for these five genes to examine differential expression between yellow and red colored bulbs. Accumulation of the DFR gene transcript only occurred in red onions. In F3 populations which originated from the cross between yellow and red parents, DFR transcript was detected only in red F3 lines, not in yellow F3 lines. To design molecular markers for selection of yellow and red DFR alleles, the DFR gene was sequenced from genomic DNA isolated from both types of onions. The genomic DNA sequence revealed the DFR gene consists of six exons and five introns. A PCR-RFLP marker was designed based on 2% polymorphic nucleotide sequence of the DFR gene between yellow and red onions. The co-segregation of markers and red color were observed in F2 segregating populations, supporting the conclusion that color difference in red and yellow onions is likely to be due to the lack of an active DFR gene.
Haejeen Bang, Angela R. Davis, Sunggil Kim, Daniel I. Leskovar, and Stephen R. King
Two loci, C and i-C, were previously reported to determine flesh colors between canary yellow and red watermelon (Citrullus lanatus). Recently, lycopene β-cyclase (LCYB) was found as a color determinant gene for canary yellow (C) and a codominant cleaved amplified polymorphic sequence (CAPS) marker was developed to identify canary yellow and red alleles. The inhibitor of canary yellow (i-C), as reported in a previous work, was not detected in our original family derived from a cross between canary yellow and red parents. To identify additional genetic determinants such as i-C, we prepared a new family using ‘Yellow Doll’ (canary yellow) and ‘Sweet Princess’ (red), which was reported to carry the inhibitor gene i-C as parents. A new distinct class of flesh color, pale yellow, was identified in the progeny from the new canary yellow × red cross. The predominant carotenoid in canary yellow and pale yellow phenotypes was neoxanthin, followed by violaxanthin and neochrome; pale yellow contained less total carotenoids, but had more minor carotenoids compared with canary yellow. The chi-square goodness-of-fit test indicated that there are two genes involved in determining flesh color among canary yellow, pale yellow, and red, but the segregation pattern did not fit the pattern as reported for an i-C gene. When the genotype of the family ‘Yellow Doll’ × ‘Sweet Princess’ was analyzed with our LCYB CAPS marker, the flesh color of every individual perfectly cosegregated with the marker. The new pale yellow phenotype also cosegregated with the marker linked to the C allele, indicating that the recessive py phenotype (pale yellow) must carry at least one of the C alleles for expression. Therefore, we propose to designate py for a pale yellow determinant along with C as a canary yellow determinant. A homozygous recessive py gene resulted in pale yellow flesh color in the presence of a dominant C.