Shoot differentiation, regeneration of cauliflower and analysis of somaclonal variation by RAPD.

Our major goal in this study was to establish and characterize an efficient callus induction and shoot regeneration protocol for cauliflower (Brassica oleracea var. botrytis). The effects of induction medium, growth regulator combination, organic component, AgNO(3), genotype and explants type on shoot differentiation on the cauliflower regeneration process were taken in account and hereby evaluated. The optimal media for shoot differentiation and rooting were modified MS medium (MS with PG-96 organic components) supplemented with NAA at 0.5 mg l(-1), TDZ at 0.25 mg l(-1), BA at 3.0 mg l(-1), AgNO(3) at 2.0 mg l(-1) and MS supplemented with IBA at 0.4 mg l(-1), respectively. Among the four varieties tested, Saixue gave the best shoot differentiation response (average over 18 shoots per explant) from the upper section of the hypocotyls. We have so far obtained over 500 regenerated plants under this improved protocol. We have further analyzed the somaclonal variation of regenerated plants at the DNA level by using the RAPD molecular markers. By PCR amplification, we were able to amplify 75 scoreable bands from 15 primers out of 40 arbitrary primers screened, where 35 of them were monomorphic and 40 polymorphic bands (53.3%) in four varieties studied. The absence of polymorphism among regenerated plants from the same variety indicated the conformity of the regeneration protocol.

Phylogenetic relationships among genotypes of worldwide collection of spring and winter ryes (Secale cereale L.) determined by RAPD-PCR markers.

Genetic similarities among 20 spring and 22 winter accessions of agronomically different ryes from fourteen countries were estimated by employing random amplified polymorphic DNA (RAPD) techniques. Cluster analysis of genetic distance data showed that 42 genotypes were readily classifiable into two main groups: spring and winter groups. Within the spring group, cultivars fell into a North European and an American-Chinese group. Cultivars of winter rye fell into four groups: Northern European, Russian, American and Chinese lines. A UPGMA-dendrogram based on genetic distances of cultivars of rye within the winter and spring groups showed that the clusters corresponded well to their geographical locations. The results indicated that isolation has played an important role in the evolution of rye, and that temporal isolation has influenced the genetic diversity of rye more than geographical isolation. In this experiment, RAPD proved to be a rapid, reliable and practicable method of revealing polymorphisms in rye populations.

Assessment of genetic variation in timothy (Phleum pratense L.) using RAPD and UP-PCR.

DNA-based fingerprinting technologies including random amplified polymorphic DNA (RAPD) and universally primed PCR (UP-PCR), a novel method for studying genetic variation, were employed as genetic markers for assessing genetic diversity and relationships in timothy (Phleum pratense L.). This study sought to identify the genetic background of the genotypes used in timothy breeding. Thirty eight genotypes from fifteen countries were used as test materials. RAPD and UP-PCR dendrograms based on 132 (from 3 primers) and 44 highly reproducible bands, respectively, were analyzed. The electrophoretic gels showed that the PCR products were informative and polymorphic. Different geographic genotype groups were distinguished according to the combined RADP and UP-PCR results. The results demonstrate that methods based on molecular fingerprinting can be used for timothy identification.