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Patrick J. Conner, Susan K. Brown, and Norman F. Weeden

Genetic linkage maps were created for three apple (Malus ×domestica Borkh.) cultivars using data from two progenies (`Wijcik McIntosh' xNY 75441-67 and `Wijcik McIntosh' xNY 75441-58). The maps consist primarily of randomly amplified polymorphic DNA (RAPD) markers, but also contain six isozyme loci and four morphological markers (Rf, fruit skin color; Vf, scab resistance; Co, columnar growth habit; Ma, malic acid). Maps were constructed using a double pseudotestcross mapping format and JoinMap mapping software. An integrated `Wijcik McIntosh' map was produced by combining marker data from both progenies into a single linkage map. Homologous linkage groups from paternal maps were paired with their counterparts in the `Wijcik McIntosh' map using locus bridges composed of markers heterozygous in both parents of a progeny. The `Wijcik McIntosh' map consists of 238 markers arranged in 19 linkage groups spanning 1206 cM. The NY 75441-67 map contains 110 markers in 16 linkage groups and the NY 75441-58 map consists of 183 markers in 18 linkage groups. The average distance between markers in the maps was ≈5.0 cM.

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Carlos A. F. Santos and Philipp W. Simon

Markers were placed on linkage groups, ordered, and merged for two unrelated F2 populations of carrot (Daucus carota L.). Included were 277 and 242 dominant Amplified fragment-length polymorphism (AFLP) markers and 10 and eight codominant markers assigned to the nine linkage groups of Brasilia × HCM and B493 × QAL F2 populations, respectively. The merged linkage groups were based on two codominant markers and 28 conserved dominant AFLP markers (based upon sequence and size) shared by both populations. The average marker spacing was 4.8 to 5.5 cM in the four parental coupling phase maps. The average marker spacing in the six merged linkage groups was 3.75 cM with maximum gaps among linkage groups ranging from 8.0 to 19.8 cM. Gaps of a similar size were observed with the linkage coupling phase maps of the parents, indicating that linkage group integration did not double the bias which comes with repulsion phase mapping. Three out of nine linkage groups of carrot were not merged due to the absence of common markers. The six merged linkage groups incorporated similar numbers of AFLP fragments from the four parents, further indicating no significant increase in bias expected with repulsion phase linkage. While other studies have merged linkage maps with shared AFLPs of similar size, this is the first report to use shared AFLPs with highly conserved sequence to merge linkage maps in carrot. The genome coverage in this study is suitable to apply quantitative trait locus analysis and to construct a cross-validated consensus map of carrot, which is an important step toward an integrated map of carrot.

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Paul Skroch, Jim Nienhuis, Geunwha Jung, and Dermot Coyne

Currently, we are studying the genetics and linkage relationships of important quantitative and qualitative traits in common bean, including disease resistances, plant architecture, seed size and shape, and pod size, shape, and fiber content. Study of the genetics of these traits is being facilitated through the use of RAPD marker-based linkage maps in four RI populations. Cultivated P.vulgaris has two primary centers of diversity—Meso-american and Andean, the RI populations used for mapping are Meso x Andean (Bat93 x Jalo EEP558 and Eagle x Puebla 152), Andean x Andean (PC50 x Xan159), and Meso x Meso (BAC6 x HT7719) crosses. Maps in these four populations are being integrated through the use of cosegregating markers. Integration of maps will allow integration of the linkage relationships of relevant genes and also allow more efficient sampling of markers for future linkage studies.

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Mark J. Bassett

Dry seeds of common bean (Phaseolus vulgaris L.) were treated with 20 krad (1 rad = 0.01 Gy) of gamma rays to induce plant mutations to be used as genetic markers in mapping studies. Four leaf mutants are described and illustrated. Inheritance studies demonstrated that each is controlled by a single recessive gene. The proposed gene symbols are: cml for chlorotic moderately lanceolate leaf, lbd for leaf-bleaching dwarf, glb for glossy bronzing leaf, and 01 for overlapping leaflets. Linkage tests involving cml and nine previously reported marker mutants failed to detect any linkages.

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Cholani K. Weebadde and James F. Hancock

While it is of great significance for strawberry breeders to know the genetics of day-neutrality (DN), evidence for inheritance of the trait is still contradictory. A linkage mapping approach is being used to determine how many QTL regulate DN and the proportion of the variability explained by each. A preliminary genetic linkage map was constructed for 125 individuals of the day neutra× short day (SD) cross `Tribute' × `Honeoye' using single dose restriction fragments (SDRFs) of amplified fragment length polymorphic (AFLP) markers. Over 500 SDRFs from 55 AFLP primer combinations were used to build the map using the software tool Join Map 3.0 at a LOD score of 3.0. Single marker analysis using WinQTL cartographer software previously determined 27 SDRF markers to co-segregate with DN for 57 individuals of the mapping population phenotyped in the field for the years 2002 and 2003, indicating putative QTL for DN. These markers were included in the linkage analysis and seven of them mapped to five different linkage groups that may indicate the quantitative nature of the trait. For determining QTL and percentage of phenotype governed by each QTL, however, accurate phenotypic evaluation is critical. Therefore, controlled environment (growth chamber) studies were used to obtain flowering response data under SD and long day (LD) conditions with two day/night temperatures. This study was conducted for the entire mapping population (over 400 individuals) so that QTL detected can be confirmed by fine mapping the QTL regions. We will also test how robust the QTL detected are, by analyzing the same segregating population at six different field locations in the United States (California, Maryland, Michigan, Minnesota, New York, and Oregon) for their flowering response under SD and LD conditions.

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Amnon Levi, C.E. Thomas, Angela Davis, O.U.K. Reddy, Y. Xu, X. Zhang, A. Hernandez, G. Gusmini, Todd C. Wehner, J. King, and S.R. King

A genetic linkage map was constructed for watermelon based on a testcross population and an F2 population. The testcross map includes 312 markers (RAPD, ISSR, AFLP, SSR, and ASRP). This map covered a genetic distance of 1385 cM, and identified 11 large (50.7-155.2 cm), five intermediate (37.5-46.2 cm), and 16 small linkage groups (4.2-31.4 cm). Most AFLP markers are clustered in two linkage regions, while all other markers are randomly dispersed throughout the genome. Many of the markers in this study were skewed from the classical (Mendelian) segregation ratio of 1:1 in the testcross or 3:1 in the F2 population. The order of the markers within linkage groups was similar in the testcross and F2 populations. Additionally, a cDNA library was constructed using RNA isolated from watermelon flesh 1 week (rapid cell division stage), 2 weeks (cell growth and storage deposition stage), 4 weeks (maturation stage), and 5 weeks (mature fruit) after pollination. More than 1020 cDNA clones were sequenced, and analyzed using the basic local alignment search Tool (BLAST). The sequenced cDNA clones were designated as expressed sequenced tag (EST). The ESTs were searched for simple sequence repeats. About 7% of the ESTs contained SSR motifs. The ESTs containing SSRs are being used to design PCR primers and the putative markers are being tested for polymorphism among the parental lines of the mapping populations. Polymorphic markers will then be mapped using the mapping populations.

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Gino E. Beltrán, Geunwha Jung, James Nienhuis, and Mark J. Bassett

The development of a complete linkage map, including both classical (visible) and molecular markers, is important to understand the genetic relationships among different traits in common bean (Phaseolus vulgaris L.). The objective of this study was to integrate classical marker genes into previously constructed molecular linkage maps in common bean. Bulked segregant analysis was used to identify 10 random amplified polymorphic DNA (RAPD) markers linked to genes for five classical marker traits: dark green savoy leaf (dgs), blue flower (blu), silvery [Latin: argentum] green pod (arg), yellow wax pod (y) and flat pod (a spontaneous mutation from round to flat pod in `Hialeah' snap bean). The genes for dark green savoy leaf (dgs) and blue flower (blu) were located in a previously constructed molecular linkage map. These results indicate that classical marker genes and molecular markers can be integrated to form a more complete and informative genetic linkage map. Most of the RAPD markers were not polymorphic in the two mapping populations used, and molecular markers from those mapping populations were not polymorphic in the F2 populations used to develop the RAPD markers. Alternative genetic hypotheses for the pod shape mutation in `Hialeah' are discussed, and the experimental difficulties of pod shape classification are described.

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Hongrun Yu and Thomas M. Davis

As part of a strawberry (Fragaria sp.) genome mapping project, we studied the linkage relationship between runnering and phosphoglucoisomerase PGI-2 allozymes in diploid strawberry. The respective r and Pgi-2 loci were found linked with a recombination frequency of 18.1% ± 1.6%(a map distance of 18.9 ± 1.6 cM). This is the second reported linkage in strawberry. The linkage between runnering and phosphoglucoisomerase allozymes, if conserved at the octoploid level, might provide a means of marker-assisted selection for the nonrunnering and bushy branching growth habits in cultivated strawberry. Severe distortion of monogenic segregation ratios was observed for runnering and PGI-2, and also for an unlinked locus for shikimate dehydrogenase allozymes. Alleles from the perpetual flowering (alpine F. vesca) parents were favored in this distortion. This phenomenon should be considered in future genetic studies using crosses between alpine and nonalpine strawberries.

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A. Levi, C. E. Thomas, A. Davis, O.U.K. Reddy, Y. Xu, X. Zhang, S. King, A. Hernandez, G. Gusmini, and T. Wehner

Genetic linkage map is being constructed for watermelon based on a testcross population and an F2 population. The testcross map comprises 262 markers (RAPD, ISSR, AFLP, SSR and ASRP markers) and covers 1,350 cM. The map comprises 11 large linkage groups (50.7–155.2 cM), 5 medium-size linkage groups (37.5–46.2 cM), and 16 small linkage groups (4.2–31.4 cM). Most AFLP markers are clustered on two linkage regions, while all other marker types are randomly dispersed on the genome. Many of the markers in this study are skewed from the classical (Mendelian) segregation ratio of1:1 in the testcross or the 3:1 ratio in the F2 population. Although the skewed segregation, marker order appeared to be consistent in linkage groups of the testcross and F2 population. A cDNA library was constructed using RNA isolated from watermelon flesh 1 week (rapid cell division stage), 2 weeks (cell growth and storage deposition stage, 4 weeks (maturation stage), and 5 weeks (postmaturation stage) post pollination. Over 1,020 cDNA clones were sequenced, and were analyzed using the Basic Local Alignment Search Tool (BLAST). The sequenced cDNA clones were designated as expressed sequenced tag (EST) markers and will be used in mapping analysis of watermelon genome.

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A. Levi, C. E. Thomas, J. Thies, A. Simmons, Y. Xu, X. Zhang, O.U.K. Reddy, A. Davis, S. King, and T. Wehner

Genetic linkage map is being constructed for watermelon based on a testcross population and an F2 population. About 51.0% and 31.8% of the markers in the testcross and F2 populations are skewed form the expected segregation ratios. AFLP markers appeared to be clustered on linkage regions, while ISSR and RAPD markers are randomly dispersed on the genome. AFLP markers also have greater genetic distances as compared with ISSR and RAPD markers, resulting in significant increase of map distance. An initial genetic map (based on the testcross population) that contains 27 ISSR and 141 RAPD markers has a total linkage distance of 1,166.2 cM. The addition of 2 ISSR, 8 RAPD and 77 AFLP markers increased the genetic distance of the map to 2,509.9 cM. Similar results with AFLP markers were also shown in mapping experiments with an F2S7 recombinant inbred line (RIL) population that was recently constructed for watermelon. Although the skewed segregation, marker order appeared to be consistent in linkage groups of the testcross and the F2 population. Experiments with SSR, and EST markers are being conducted to saturate the linkage map of watermelon genome.