Dominant Molecular Markers Research Articles

Evolutionary relationships within recently radiated taxa: comments on methodology and analysis of inter‐simple sequence repeat data and other hypervariable, dominant markers

AbstractHypervariable, dominant molecular markers have become more common as characters for reconstructing evolutionary relationships, largely due to the difficulty in locating useful variation in DNA sequences in some very recently radiated taxa. Several of the methodological and analytical issues relating to these types of data are discussed, specifically focusing on inter‐simple sequence repeat (ISSR) data. Caution in avoiding redundancy in primer selection is recommended, as well as the use of 3' anchored primers. Manual versus automated methods are briefly discussed, in addition to the influence of fluorescent markers on ISSR PCR reactions. A series of clustering and phylogenetic analysis methods are compared. Several similarity coefficients are available for clustering analyses of ISSR data. Those that exclude shared‐absence characters appear most appropriate, such as Nei & Li's (1979) coefficient. However, more than one coefficient with this title is accessible from different software programs; care should be taken to be certain of the characteristics of a given coefficient. The potential suitability of Dollo, Fitch, or Camin‐Sokal Parsimony for ISSR analyses is also discussed.

Genetic Models of Host–pathogen Gene Interaction Based on Inoculation of Loblolly Pine Seedlings with the Fusiform Rust Fungus

Loblolly pine (Pinus taeda) seedlings from 12 full-sib families obtained from a six-parent half diallel mating design were challenged in a greenhouse using two basidiospore inocula of the fusiform rust fungus (Cronartium quercuum f.sp. fusiforme) at extremely high spore density. Each basidiospore inoculum originated from a mixed gall collection of aeciospores obtained from field-infected trees. Assessments at 4.5 months after inoculation showed that rust disease levels were high for every full-sib family and were typically above 90% for most full-sib families for both inocula. However, disease (% galled) for family E by A progeny, even at 9 months post inoculation, was lower, around 75%. A genetic model for interaction of two pairs of genes was proposed to explain the observed infection levels (% galled) in this diallel based on a gene-for-gene hypothesis. The putative genotypes of host parents and virulence compositions of mixed inocula were postulated. A bulk-segregant analysis approach based on phenotype (gall vs. no gall) was used to search for dominant molecular markers associated with the potential resistance genes in the host parents. A few candidate marker polymorphisms were observed between the gall vs. no gall bulks; however, none of the candidates appropriately co-segregated with phenotype when tested across the progeny set. An alternative model involving recessive resistance controlled by a single locus was also considered, but as with the two gene model, no markers to support the appropriateness of the recessive resistance model were observed.

An Overview of RAPD Analysis to Estimate Genetic Relationships in Lowbush Blueberry

SUMMARY Randomly amplified polymorphic DNA (RAPD) analysis, a simple dominant molecular marker technique, has been used extensively for cultivar identification and relatedness studies in many perennial woody species. Thus, this technique should provide genetic information for lowbush blueberry (Vaccinium angustifolium Ait.). Young leaves of lowbush blueberry from field clones with varying phenotype were collected for DNA extraction. Pre-screening of RAPD primers resulted in 11 polymorphic primers and 140 consistent RAPD fragments. Eight primers were selected as useful for our study, the fragments scored and the data analyzed with Genstat5 to calculate similarity, produce dendrograms and perform a principal coordinate analysis. The RAPD analysis was able to identify distinct field clones. Average genetic similarity among field clones was 68% reflecting expected genetic variation. Approximately 15% of the field clones were not related. RAPD analysis is a useful tool for genetic relationship studies in lowbu...

Identification of Random Amplified Polymorphic DNA and Simple Sequence Repeat Markers Linked to Powdery Mildew Resistance in Common Wheat Cultivar Brock

A total of 350 rapid amplified polymorphic DNA (RAPD) primers and 100 simple sequence repeat (SSR) primer pairs were screened to identify polymorphic markers associated with powdery mildew resistance. Only primer OPP15 produced a 900bp reproducible DNA fragment (OPP15900) in the resistant parent cv. Brock and most of the resistant individuals, but this DNA fragment was absent in susceptible parent Jing411 and Line 015. The progeny, including 218 resistant and 81susceptible lines, derived from a cross Line 015/Brock//Jing4112 was used for linkage analysis. 209 resistant and 8 susceptible individuals yielded OPP15900 products, but 73 susceptible and 9 resistant ones yielded no OPP15900 products. One dominant RAPD molecular marker OPP15900 linked to powdery mildew resistance gene was identified in Brock with a genetic distance of 6.0 cM. A SSR marker Xgwm114 was also proved to link with the powdery mildew resistance and genetic distance of 9.3 cM. These two new molecular markers are useful for facilitating selection and pyramiding the resistance genes in wheat breeding.

Paternity analysis in the Antarctic brooding sea urchin Abatus nimrodi . A pilot study

The genus Abatus (Echinoidea: Spatangoida: Schizasteridae), endemic to the Southern Ocean, consists of several species which, like many other Antarctic marine invertebrates, brood their offspring. The modality of fertilization is not known in these species, whose direct observation and sampling are difficult. Parentage analyses by means of molecular markers may help to gain information on this stage of the life-cycle. In this pilot study, we analysed a brooding female and her offspring with dominant molecular markers—RAPD (Random Amplified Polymorphic DNA). We established that each cohort present in the brooding pouches, i.e. gastrulae and juveniles, originated from at least two distinct father genotypes. Our original method of analysis of dominant marker data should have vast applications since it allows one to test, not only (1) the hypothesis that the progeny of a given mother originates from a single father, but also (2) the temporal stability of the paternal gene pool.

Experimental population design for estimation of dominant molecular marker effect on egg‐production traits

A potential limitation of the use of a dominant molecular marker system such as DNA fingerprinting (DFP) is the inability to distinguish homozygous from heterozygous allele state in an individual, and a resulting inaccuracy in estimating effects of the marker alleles. The objective of this study was to accurately estimate the effect of DFP markers on egg-production traits. A BC1 population was produced from two distinct layer lines. Four DFP bands, each originating predominantly in one of the two parental lines, were evaluated for linkage with egg-production quantitative trait loci in the BC1 population. The egg-production traits consisted of eight early period and seven late period measurements. Eight marker-trait linkages were identified out of 60 total statistical tests. By utilizing information on frequency of DFP bands in two parental lines, selecting F1 sires with DFP bands present, and backcrossing to the line lacking these bands, the population design allowed definitive identification of the DFP zygosity in the BC1 resource population hens. In this manner, accurate estimates of marker allele effects on egg-production traits were obtained from the dominant marker system of DNA fingerprinting.

Multidrug resistance as a dominant molecular marker in transformation of wine yeast

Pure wine yeast cultures are increasingly used in winemaking to perform controlled fermentations and produce wine of reproducible quality. For the genetic manipulation of natural wine yeast strains dominant selective markers are obviously useful. Here we demonstrate the successful use of the mutated PDR3 gene as a dominant molecular marker for the selection of transformants of prototrophic wine yeast Saccharomyces cerevisiae. The selected transformants displayed a multidrug resistance phenotype that was resistant to strobilurin derivatives and azoles used to control pathogenic fungi in agriculture and medicine, respectively. Random amplification of DNA sequences and electrophoretic karyotyping of the host and transformed strains after microvinification experiments resulted in the same gel electrophoresis patterns. The chemical and sensory analysis of experimental wines proved that the used transformants preserved all their useful winemaking properties indicating that the pdr3-9 allele does not deteriorate the technological properties of the transformed wine yeast strain.

Construction of a genetic linkage map in tetraploid species using molecular markers.

This article presents methodology for the construction of a linkage map in an autotetraploid species, using either codominant or dominant molecular markers scored on two parents and their full-sib progeny. The steps of the analysis are as follows: identification of parental genotypes from the parental and offspring phenotypes; testing for independent segregation of markers; partition of markers into linkage groups using cluster analysis; maximum-likelihood estimation of the phase, recombination frequency, and LOD score for all pairs of markers in the same linkage group using the EM algorithm; ordering the markers and estimating distances between them; and reconstructing their linkage phases. The information from different marker configurations about the recombination frequency is examined and found to vary considerably, depending on the number of different alleles, the number of alleles shared by the parents, and the phase of the markers. The methods are applied to a simulated data set and to a small set of SSR and AFLP markers scored in a full-sib population of tetraploid potato.

Predicting parental genotypes and gene segregation for tetrasomic inheritance

Recent genome mapping projects in tetraploid plant species require a method for analysing the segregation patterns of molecular marker loci in these species. The present study presents a theoretical model and a statistical analysis for predicting the genotypes of a pair of tetraploid parents at a codominant (for example, RFLPs, microsatellites) or dominant (for example, AFLPs, RAPDs) molecular marker locus based on their and their progeny’s phenotypes scored at that locus (gel-band patterns). The theory allows for null alleles and for any degree of double-reduction to be modelled. A simulation study was performed to investigate the properties of the theoretical model. This showed that in many circumstances both the parental genotypes can be correctly identified with a probability of nearly 1, even when the molecular data were complicated by null alleles or double-reduction. Configurations where the parental genotype cannot be identified are discussed. The power to detect double-reduction varies considerably, depending on the proportion of identical alleles carried and shared by the parents, and the number of null alleles. Incorrect deductions of the occurrence of double-reduction were rare. The method was applied to data on a microsatellite locus segregating in the parents and 74 offspring of a tetraploid potato cross. Twentyfour parental configurations were consistent with the parental gel pattern, but only one of these was compatible with all the phenotypic data on the offspring. The feasibility for extending the present model to predict segregation of several linked loci, and particularly the linkage phase, is briefly discussed.

Trifluoroleucine resistance as a dominant molecular marker in transformation of strains of Saccharomyces cerevisiae isolated from wine

The resistance to 5,5,5-trifluoro- DL-leucine, encoded by the dominant allele LEU4-1, was used as a selectable marker to transform laboratory and natural Saccharomyces cerevisiae strains by the lithium acetate procedure. Results of transformation of S. cerevisiae laboratory and wine natural strains showed that trifluoroleucine resistance is a very effective selection marker and can be widely used to transform prototrophic S. cerevisiae strains. The LEU4-1 gene could also be exploited to improve wine flavour, as indicated by the higher isoamyl alcohol content of the transformants compared to the parental strains.

Trifluoroleucine resistance as a dominant molecular marker in transformation of strains of Saccharomyces cerevisiae isolated from wine.

The resistance to 5,5,5-trifluoro-DL-leucine, encoded by the dominant allele LEU4-1, was used as a selectable marker to transform laboratory and natural Saccharomyces cerevisiae strains by the lithium acetate procedure. Results of transformation of S. cerevisiae laboratory and wine natural strains showed that trifluoroleucine resistance is a very effective selection marker and can be widely used to transform prototrophic S. cerevisiae strains. The LEU4-1 gene could also be exploited to improve wine flavour, as indicated by the higher isoamyl alcohol content of the transformants compared to the parental strains.

Estimation of the position effect and action mode of a semi-lethal factor locus on a DNA polymorphism linkage map in silkworm, Bombyx mori.

Distorted segregation has been repeatedly observed in molecular marker linkage mapping of various plants and animals where distant crosses were made. It may be caused by a semi-lethal factor acting in the filial generations. Although general methods have been developed to find semi-lethal factors, so far no method is available to estimate semi-lethal factors in the organisms, for example Bombyx mori and Drosophila melanogaster, in which no crossing-over occurs in meiosis in either of the sexes. In this report, a method is presented for estimating the recombination values between a semi-lethal factor locus and neighboring dominant molecular markers, e.g. RAPDs and AFLPs, and the relative viability of gametes or zygotes affected by the semi-lethal factor in an F2 population for such species by using the maximum likelihood method associated with the expectation maximization (EM) algorithm. Five selection models of gamete or zygote were considered, and the most likely one was determined by goodness of fit of the observed counts of phenotypes to the expected ones under the models. The method was applied to segregation data of RAPD markers of an F2 population with 101 individuals developed from the `C108' × `p50' cross in silkworm, B. mori. The presence of two semi-lethal factor loci (L 1 and L2) located on the linkage groups 22a and 27b causing partial selection was suggested. The semi-lethal factors L1 and L2 were theoretically predicted to affect viability of male gametes and zygotes, respectively.