Our objective was to identify QTL for seed weight (SW), length, and height segregating in a recombinant inbred line (RIL) population from the cross `PC-50' (Larger SW) × XAN-159 (Smaller SW). The parents and RILs were grown in two separate greenhouse experiments in Nebraska, and in field plots in the Dominican Republic and Wisconsin. Data analysis was done for individual environments separately and on the mean over all environments. A simple linear regression analysis of all data indicated that most QTL appeared to be detected in the mean environment. Composite interval mapping (CIM) analysis was then applied to the means over environments. Eight QTL for SW were detected on common bean linkage groups (LGs) 3, 4, 5, 6, 7, and 8. All eight markers associated with these QTL were significant in a multiple regression analysis (MRA), where the full model explained 63% of the variation among SW means. Six QTL for seed length were detected on LGs 2, 3, 4, 8, and 11 using CIM. The markers associated with the three seed length QTL on LGs 2, 8, and 11 were significant in a MRA with the full model explaining 48% of the variation among seed length means. Three QTL for seed height on LGs 4, 6, and 11 explained 36% of the phenotypic variation for trait means. Four of the six QTL for seed length and two of three QTL for seed height also appeared to correspond to QTL for SW.
Soon O. Park, Dermot P. Coyne, Geunhwa Jung, Paul W. Skroch, E. Arnaud-Santana, and James R. Steadman
Christopher M. Long, Colleen A. Mulinix, and Amy F. Iezzoni
Microspore-derived callus cultures were obtained by anther culture of `Emperor Francis' sweet cherry (Prunus avium L.). Branches were removed from the field in January and March and forced in the laboratory. When the microspores reached the uninucleate stage, anthers were placed on modified Quoirin and Lepoivre liquid culture medium containing 4.4 μm BA and 4.5 μm 2,4-D. After ≈60 days, callus that emerged from the anthers was placed on woody plant medium supplemented with 1 μm 2,4-D and 3 μm 2iP and routinely transferred. The resulting 270 callus cultures were screened for two allozymes heterozygous in `Emperor Francis', Pgi-2 and 6-Pgd-1. Of the 270 callus cultures, 154 expressed only one allele each for Pgi-2 and 6-Pgd-1; thus, they were considered microspore-derived. The microspore-derived callus cultures can be used as a linkage mapping population. Chemical names used: 6-benzyladenine (BA); 2,4-dichlorophenoxyacetic acid (2,4-D); N6-(2-isopentenyl)-adenine (2iP).
Catherine M. Ronning and Raymond J. Schneg
While Random Amplified Polymorphic DNA (KAPD) has been used successfully in genetic analysis of several crop plants, the method poses difficulties with a heterozyaous species such as Theobroma cacao due to the dominant phenotypic expression of bands. A backcross family of the cultivars `Catoneo' and `Pound 12' was analyzed to determine the efficacy of this technique in analyzing cacao populations. A primary screen of the parents and F1 was conducted with 180 KAPD primers; of these, 39.5% either-did not amplify or did so poorly or irreproducibly, while 60.5% amplified well. Phenotypes produced by 42 primers represented possible test crosses, which can be used in linkage mapping. Genomic DNA from 50 individuals of the backcross population were then amplified with these 42 primers, which in most cases resulted in 1:1 segregation of bands. Preliminary experiments show that the Stoffel fragment of Taq DNA polymerase may provide additional markers. These results indicate that it should be possible to use RAPD bands as molecular markers to study the cacao genome.
John A. Juvik
Investigating the chemical constituents that determine human preferences for cooked vegetable flavor and aroma is complicated by experimental limitations. Several to many biochemicals interact with each other and with textural properties to influence perception of eating quality. This is particularly true for volatile compounds associated with aroma, where differences in concentration, volatility, reactivity, chemical stability, thresholds of perception, and duration of receptor bonding generate transient stimuli that are integrated into the sensory evaluation of quality. This paper describes methodology that can isolate, identify, and quantify the effect of chemical constituents that influence flavor and aroma using populations segregating for genes controlling eating quality. A F2:3 population derived from a cross between two sweet corn inbreds that differed in kernel characteristics associated with eating quality were assayed for variation in chemical, physical, and sensory characteristics. Because most aromatic constituents of sweet corn are generated during cooking, kernel tissue samples were autoclaved and analyzed by gas chromatography. Panel variation in sample preference were found to be controlled by three overlying factors—taste, texture, and aroma—the relative importance of each being 45.1%, 30.5%, and 24.4%, respectively. DNA marker technology was employed to generate a linkage map of this population that was sufficiently saturated with probes to allow for the identification and mapping of genes controlling each characteristic. This information improves selection methodology in a breeding program aimed to develop germplasm with superior eating quality.
Paul Skroth, Jim Nienhuis, Geunhwa Jung, and Dermont Coyne
Knowledge of genetic relationships and genetic diversity among accessions is essential for the efficient construction, maintainance and utilization of large ex-situ germplasm collections. Furthermore, streamlining of large collections into care collections necessitates validation of germplasm sampling techniques. DNA molecular markers provide potentially unbiased estimators of genome diversity end may facilitate organization, maintainance, and sampling of plant genetic resources. Our data suggests that RAPD markers will be o good tool for testing tore collection concepts and organizing genetic diversity in common bean. However, the genomic distribution of markers is unknown. Currently we are using recombinant inbred (RI) populations to place RAPD markers in the context of the bean genetic map. We hove evaluated the the distribution of RAPD markers in three RI populations: Bat93 × Jalo EEP558, PC50 × Xan159, and BAC6 × HT7719. Cultivated P.vulgaris has two primary renters of diversity Mesoamerican and Andean, the RI populations used for mapping RAPD markers ore Meso × Andean, Andean × Andean, and Meso × Meso crosses respectively. In the Bat93 × Jalo EEP558 population 383 markers have been mapped for a map length of 735 cM. However, approximately 150 of these markers ore members of 9 dusters which span only 90 cM. This inter gone pool mop is being integrated with linkage mops constructed in the other two populations to compare within and between gene pool marker distributions and to evaluate clustering of markers on the different mops. Implications for the application of RAPD markers will be discussed.
Zhan'ao Deng, Fred G. Gmitter Jr., Shunyuan Xiao, and Shu Huang
Citrus tristiza virus (CTV) is the most-significant viral pathogen of citrus in the world. Rapid decline of trees on sour orange and stem pitting of grapefruit and sweet orange, two diseases induced by CTV, severely jeopardize citrus production worldwide. It is recognized that all future rootstocks should be resistant to this virus, and scion resistance to stem pitting stains is desirable. To facilitate introgression of the CTV resistance gene from Poncirus trifoliata and development of CTV-resistant varieties in citrus, gene mapping projects have been initiated and more than a dozen RAPD markers have been identified with tight linkage to the resistance gene. As part of our efforts to use marker-assisted selection with a large number of crosses, and ultimately to accomplish map-based cloning of the CTV resistance gene, we have been converting the most tightly linked RAPD markers into SCAR (sequence characterized amplified region) markers by cloning, sequencing the marker fragments, and designing locus-specific primers. One codominant and several dominant SCARs have been developed thus far. The updated progress and utilization of these SCARs in marker-assisted selection and possibly in characterization of a BAC library will be presented and discussed.
James M. Bradeen and Philipp Simon
The Y2 locus conditions α- and β-carotene accumulation in the xylem (core) of carrot roots. The dominant allele suppresses carotene, but not xanthophyll accumulation, resulting in yellow-cored roots. Individuals homozygous for the recessive allele are rich in carotenes and are therefore orange-cored. Increased consumer interest in high carotene produce requires improved understanding of carotene biosynthesis and color development and more-efficient breeding techniques. We examined 103 F2 individuals generated from inbred populations differing in core carotene content. Bulked segregant analysis identified AFLP bands putatively linked to Y2. Linkage was confirmed for some bands by mapping. Linked bands were excised from gels, re-amplified, cloned into pGEM vectors, and sequenced. Cloned fragments and sequence information were used to characterize larger genomic regions to identify codominant markers. Currently we are developing codominant, PCR-based markers that can be used to rapidly genotype individuals in breeding programs, to characterize wild, feral, and cultivated populations for diversity and evolution studies, and to examine the role of Y2 in carotene accumulation.
Eileen Kabelka, Wencai Yang, and David M. Francis
An inbred backcross (IBC) population derived from Lycopersicon hirsutum LA407 and L. esculentum was evaluated in replicated field trials to assess its potential for the improvement of red-fruited tomatoes. Significant phenotypic variation among genotypes was detected for the hue (tint), L (darkness), and chroma (saturation) of color. Significant effects due to environment and genotype × environment interactions also were observed. One superior inbred backcross line from this population, IBL 2349, was used to develop an F2 population and to explore the genetic basis of color. Two independent L. esculentum quantitative trait loci (QTL) associated with improved color were identified based on linkage to markers mapping to chromosome 4 and chromosome 11. Epistatic interactions were identified between the two L. esculentum loci. Unexpected epistatic interactions also were identified between L. esculentum loci and an LA407 introgression on chromosome 7 present within IBL 2349. The two L. esculentum QTL and the epistatic interactions were confirmed in replicated trials with F3 and F4 families. The loci identified in this study and their epistatic interactions may provide additional tools for the improvement of red-fruited tomatoes in breeding programs.
Amnon Levi, Angela Davis, Pat Wechter, Alvaro Hernandez, and Jyothi Thimmapuram
A cDNA library was assembled using mRNA of watermelon fruit. The cDNA library was normalized and subtracted by hybridization with leaf cDNA of the same watermelon cultivar (Illini Red). 1,046 cDNA clones were sequenced to identify genes associated with fruit development and quality. Of 1,046 cDNA clones sequenced, 832 were unique sequences and designated as expressed sequenced tags (ESTs). Of the 832 ESTs, 205 (24.6%) have not been reported in any other plant species. Additionally, 186 ESTs (22.4%) correspond to genes with unknown function, while 441 ESTs (53.0%) correspond to genes with known function in other plant species. These ESTs are mainly associated with primary metabolism, membrane transport, cytoskeleton synthesis and structure, cell wall and cell division, signal transduction, nucleic acid binding and transcription factors, and defense and stress response. Differential expression of the ESTs was examined using microarray analysis. About 200 (24%) of the 832 ESTs showed differential expression during the development and ripening of watermelon fruit. The ESTs were also screened for simple sequence repeat (SSR) motifs. Of 832 ESTs screened, 177 contain SSR motifs. Primer pairs are being designed for these ESTs, and will be used for development of EST-SSR markers and for mapping on a genetic linkage map constructed for watermelon. This study provides valuable information on genes controlling watermelon fruit development and quality.
Rebecca Nelson Brown and James R. Myers
., for providing seed of the parental lines, and greenhouse space for the development of the mapping population, and to H.S. Paris of Newe Ya'ar Research Station, Ramat Yishay, Israel, for fruitful discussion of the genetics of fruit color in Cucurbita .