| The species of Porphyra(Bangiacea, Rhodophyta), distributed in the cold, temperate and subtropic intertide waters, are of economic importance in sea-weed farming industry. With the development of laver farming industry, those requirements for favorite lines and efficient farming techniques are more and more emphasized. However, the traditional seedling breeding of laver is mainly based on wild species without any selective breeding and genetic modification, which may deduce the adaptability to environment and resistance to disease, and may influence or even pollute the gene pool of natural species surrounded. Therefore, it’s of significance to analyze the genetic variation, to detect the specific markers for lines and stains of Porphyra, so as to reveal their genetic background, based on which, the rational strategy for seedling production can be framed and the measures to protect the laver genetic resources can be taken.Porphyra yezoensis is an ideal material for genetics and molecular biology research. Recently it has been suggested as a model plant in marine biological sciences for basic and applied studies because of its unique dimorphic generation cycle. AFLP(Amplified Fragment Length Polymorphism) reaction system was established with a breed of Porphyra thallus, and the reaction factors have been optimized. A appropriated reaction system of AFLP has been founded for Porphyra, By choosing the selection of experimental materials, extraction of genomic DNA, digestion time and dilution Times., a appropriated reaction system has been established for Porphyra thallus. Experimental materials from Qingdao beach1and beach2was used as experimental comparison. Improved CTAB methods has been used to exact DNA, and Grinding in liquid nitrogen, Adding quartz sands and Increase in cracking time were used in the experiment.3,5,7-hour digestion time was selected as a control,5h was the best time by agarose gel electrophoresis and spectrophotometric detection.5times,10times,20times,30times and40times dilution was selected in multiples, and40times was the best dilution times by agarose gel electrophoresis, spectrophotometry and denaturing polyacrylamide gel electrophoresis detection. Ligation reaction was under16℃, E+A and M+C primers were used in Pre-expanding and B1, B2and B4primers were used in selected expanding, and then Polyacrylamide gel was colored with silver-staining method, An appropriated AFLP reaction system for Porphyra thallus will lay the foundation for further study in molecular biology of Porphyra.Amplified fragment length polymorphic technique was used to analyze the genomic DNA polymorphism of13lines of Porphyra yezoensis.11of the80AFLP primer pairs gave reproducible polymorphic DNA amplification patterns, and619bands are obtained. The amplified bands ranged from40to77per primer pair shows the genetic diversity among these samples.609of AFLP bands were polymorphic loci. The proportion of polymorphic loci was98.38%. The genetic distance (D) and similarity coefficients (GS) were calculated based on the polymorphic data. UPGMA cluster indicate that samples from Haian and Ershi were with high similarity. The GS between these samples was0.595-0.845. The result showed that the genetic diversity level in Porphyra yezoensis was fairly high. Similar increasing trend was found in genetic distance coefficient and geographical distance. UPGMA cluster indicated that Porphyra yezoensis in our country was miscellaneous. |
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