Construction Of Molecular Genetic Linkage Map Of Stevia Rebaudiana

Stevia rebaudiana is one of the small economic crops. Its molecular biology research is still in the early time. This paper mainly did research onthe molecular genetic map construction in S. rebaudiana, that would lay the foundation for the QTL(quantitative trait loci)localization, gene localization and cloning, genomics research and marker-assisted selection breeding of S. rebaudiana. Specific results were as follows:Owing to the little information on the molecular markers which are needed to construct S. rebaudiana's genetic map, the molecular markers were analyzed and developed based its transcription and genome information. In present research, the EST-SSR was screened from the existing S. rebaudiana ESTs database and its distribution characteristics were analyzed. Then, the EST-SSR functional markers were expanded for exploitation and utilization. By analyzing 5573 non-redundant S. rebaudiana ESTs comprising 3.75 Mb,285 SSRs were identified. There was on the average of one SSR for every 13.2 kb ESTs. The 285 SSRs consisted of 56.5% dinucleotide,43.2% trinucleotide and 0.3% tetranucleotide repeats. Dinucleotide repeats were the predominant form in frequency. 58 pairs of primers were designed and developed from the SSR containing EST sequences. In a previous PCR analysis of the SRAP marker, this experiment established and optimized the S. rebaudiana SRAP-PCR reaction system at the dNTPs concentration of 0.2 mmol/L, Tap enzyme concentration of 0.5 U/μl and DNA concentration of 1.2μmol/L. Nineteen primers were designed for the upstream and downstream primers, forming 361 pairs of SRAP primer combinations.A 410 F1 separate population was constructed using two inbred lines of S. rebaudiana. After the screening and analysis of EST-SSR and SSR markers for polymorphism in parents and F1, two genetic maps that refered to as big leaf and small leaf material with a total length of 1215.2cM and 647.8cM were constructed, respectively. The big leaf material map contains 13 linkage groups, a total of 74 markers and the average distance between markers was 16.4cM. The small leaf material map also contains 12 linkage groups, a total of 49 markers and the average distance between markers was 13.2cM. The estimated genome size of S. rebaudiana was 2286.5cM, with the genome coverage ratios of the big leaf and small leaf materials being 53.1% and 28.3%, respectively.