Comparison of Isozyme, RAPD and AFLP Markers in Genetic Similarity Assessment of CMS Ogura F1 Hybrids of Winter Oilseed Rape (Brassica Napus L.) Parental Lines

Abstract

Isozyme, RAPD and AFLP markers were evaluated and compared for their ability to determine genetic similarity in a set of 18 parental lines of winter oilseed rape F 1 hybrids developed using CMS ogura. Five isozyme systems, 64 RAPD starters and 23 EcoRI+3/MseI+3 AFLP primer combinations generated 597 polymorphic markers. These polymorphic fragments were chosen for estimation of genetic similarity. Of the total number of polymorphic products, polymorphic zymograms constituted only 3.0% of the markers, 57 RAPD primers 37.7%, and 23 AFLP primer combinations 59.3%. The size of RAPD polymorphic products ranged from 564 to 2100 bp. On average there were four amplified bands per primer, with 61.0% polymorphism. The AFLP polymorphic fragments ranged from 72 to 1352 bp in size. AFLP assays generated 15 bands per primer pair on average and detected roughly four times more bands than with RAPD analysis. The genetic similarity coefficients based on all marker data range from 0.52 to 0.84. The correlation of genetic similarities based on RAPD and AFLP markers was 0.58. Estimated genetic similarity based on isozyme data was not correlated with genetic similarity derived from the two DNA-based markers. The dendrogram constructed with the three types of markers taken together grouped all the winter oilseed rape parental lines into several similar clusters. The genomic distribution and frequency of the RAPD and AFLP markers can serve well as estimators of genetic similarity between parental lines of F 1 CMS ogura hybrids.K Ke ey y w wo or rd ds s: : Winter oilseed rape, Brassica napus, F 1 hybrids CMS ogura, genetic similarity, molecular markers.

Assessment of Genetic Relationships in Breeding Lines and Cultivars of Brassica napus and Their Implications for Breeding Winter Oilseed Rape

“…The integrity and concentration of the DNA was confirmed by 0.8% agarose gel electrophoresis for 1h at 100 volts in Table 1. The primer combinations were selected from the linkage map for rapeseed described by Lombard and Delourme (2001) and the results obtained in the analyses of collections of different WOSR genotypes (Liersch et al 2012(Liersch et al , 2013. The AFLP procedure was performed using the standard method in accordance with the manufacturer's instructions (AFLP Plant Mapping, Applied Biosystems) and as previously described by Vos et al (1995).…”

“…The potential of isozyme markers is limited by their low frequency in different plants in comparison with markers based on DNA analysis with a theoretically unlimited number of polymorphic markers. The usefulness of random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) markers was examined to determine genetic distance between Polish winter oilseed rape breeding materials (Liersch et al 2013). Amplified fragment length polymorphism markers were used to estimate the genetic distance of a collection of 78 genotypes including male sterile and restorer lines for CMS ogura, a few B. napus cultivars, and genotypes of European and Asian origin (Liersch et al 2012).…”

“…This technique is a highly reproducible method with low error rates ( Jones et al 1997;Powell et al 1996). Amplified fragment length polymorphism markers have been widely used in many plant species to construct genetic maps, to study genetic diversity phylogenic relations and crop evolution, to evaluate germplasm, and to reveal differences between varieties and genotypes (Lübberstedt et al, 2000;Lombard and Delourme, 2001;Hartings et al, 2008;Pawłowicz et al, 2008;Dobrzycka et al, 2009;Bocianowski et al, 2011;Kozak et al, 2011;Liersch et al, 2013;Würschum et al, 2013). Lombard et al (2000), Negi et al (2004), and Faltusová et al (2011) reported that AFLP markers demonstrated the high efficiency of the AFLP technique for genome analysis in Brassica.…”

“…EcoRI primers were labeled with fluorescent dye: FAM (blue), JOE (green), or NED (yellow) (AFLP Selective Amplification Primers, Applied Biosystems). The primer combinations were selected from the linkage map for rapeseed described by Lombard and Delourme (2001) and the results obtained in the analyses of collections of different WOSR genotypes (Liersch et al 2012(Liersch et al , 2013. All polymerase chain reactions (PCR) were performed in an Eppendorf Mastercycler Gradient thermo cycler.…”

Genetic variability of cultivated Chaenomeles speciosa (Sweet) Nakai based on AFLP analysis

“…Moreover, AFLP markers provide a broader polymorphism spectrum (highest number of segments of DNA are analyzed) than the random amplified polymorphic DNA (RAPD) and isozymes markers (Liersch et al, 2013). These characteristics of AFLP aid in evaluating genetic relationships among cultivars and identifying cultivars of many crops (Ude et al, 2003;Raji et al, 2009;Faria-Tavares et al, 2013).…”