| Porphyra yezoensis(P. yezoensis) is one of the most important economically marine algae. In recent years, the quality of the P. yesoensis is reduced, which greatly affects the development of seaweed industry. The cultivation of best quality algae with stress resistance becomes the urgent need. Germplasm resource of high quality is the key point of breeding stress resistance species. However, the research on the breeding of marine algae started late, and the basic research of it is weak. At present, there is no physiological and molecular markers to screen the excellent germplasm resources. Therefore, it is important to find the key position of P. yesoensis in response to abiotic stress and develop the molecular markers. In this study, we focused on resistant breeding of P. yezoensis. More attentions were paid to determine the key position of resistance characteristics under salt stress in P. yezoensis and develop SSR markers for resistance characteristics of quantitative trait loci. In addition, the genetic diversity was extended by artificial mutation, all of which laid the foundation for the quantitative trait loci of the resistance traits. These results mainly include the follows:(1) The photosynthetic(PS) responses of P. yezoensis blades under salt stress were studied. Our results showed that when the effective photochemical quantum yield of PSII almost decreased to zero under high salt stress, while YI still had a relative high activity rate. PSII is therefore more sensitive to salt stress than PSI. These results demonstrated that under severe salt stress(120‰ salinity), the electro source of PSI was derived from a source other than PSII, and that stromal reductants probably supported the operation of cycle electron flow around PSI. Under salt stress, the starch content decreased and the soluble sugar levels increased. The G6 PDH and 6PGDH(EC 1.1.1.44) activities increased, but the GAPDH activity decreased. Furthermore, the NADPH content increased, and NADH decreased, which suggested that soluble sugar entered the oxidative pentose phosphate pathway(OPPP). All these results suggested that NADPH from OPPP increases the cyclic electron flow around PSI in high intertidal macroalgae under severe salt stress.(2) Based on the sequenced genomic, 700 SSR markers of complete repeat were developed. There are 254495 SSR sites in the 270 M genomic, with one SSR marker every 1 kb on average. These results showed that the SSR markers were abundant, including all the 2-6 bases SSR makers. Among the genomic two base repeats have the largest proportion, followed by three bases. The highest proportion of repeat types is CG/GC type, followed by CGG/CCG type. The polymorphism of 700 SSR markers was analyzed in 16 P. yezoensis conchocelis from different places. The results demonstrated that 98 SSR markers were polymorphic in these samples. Genetic analysis was carried out on the polymorphic makers according to the PCR results. By GenALEx6.3 software: The average value of alleles was 2.479, effective alleles(Ne) was 1.558, diversity information index(I) was 0.543, observed heterozygosity(Ho) was 0.168, expected heterozygosity(He) was 0.32, PIC was 0.35. Cluster analysis was carried out by UPGMA method. The results showed that these 16 conchocelis of P. yezoensis were clustered into two main branches. All these results suggested that the gerplasm of P. yezoensis have been mixed, and there is gene flow exchange between them.(3) In this study, the EMS mutation and heavy ion beam were used to get the mutant strains of P. yezoensis conchocelis. Research shows that the treatment of 0.2 M EMS concentration for 2 h, 0.4 M EMS concentration for 1.5 h and 0.6 M EMS concentration for 1 h are effective ways to get P. yezoensis mutant strains. Which provides a reference for other uses of EMS induction in P. yezoensis. Heavy ion beam irradiation method showed that the 100-150 GY dose was effective dose. Twenty conchocelis were randomly selected from the mutant strains, then the genetic diversity was analyzed by SSR markers. The average value of alleles(Na) was 2.842, effective alleles(Ne) was 1.935, diversity information index(I) was 0.756, observed heterozygosity(Ho) was 0.656, expected heterozygosity(He) was 0.464, and the PIC was 0.39. The samples were clustered approximately in accordance with the dose of heavy ion beam, and divided into several sub categories which were clustered with the control conchocelis. The genetic diversity of these mutant strains is more abundant then that of the16 P. yezoensis conchocelis. overall, the mutagenic method can enrich the genetic diversity of P. yezoensis. |
