Pure Line Cultivation Of Porphyra Haitanensis And The Primary Research Of Microsatellite Marker Test

The release of free-1 iving vegetative eel 1 were successful ly achieved by enzymolysi.de of Porphyra haitanensis thallus, the enzyme was extracted from snail. Also the morphogenesis of single somatic cell was described. After enzymolysis, single somatic cell was picked out under inverted microscope, and incubated in 96 microplate. The living cells grew into conchocelis, blade and callus through different developing routes. Besides, by the comparion of size and germination fashion, sexual cell can be distinguished from somatic cell, that' s important for the achievement of pure line conchocelis establishment.The quantity of conchocelis was multiplied using suspension culture, and further cultured as the same as the traditional mariculture. Comparison of thallus of pure line with the control group have revealed some superior qualities.Taking the advantage of microsatellite marker in individual identification, plan to verificate the blades of pure line on molecular level. Fisrt stage is to obtain useful microsatellite markers.From Porphyra yezoensis ESTs ,all dinucleotide repeats with n>7 and all possible trinucleotide reeats with n > 5 were extracted. Total 211 non-redundant microsatellites were isolated from 20,979 ESTs sequences, amount to 1. 01% of the population of ESTs in the database. Among them , 16. 6%, 35 dinucleotide repat and 83. 4%, 176 trinucleotide repeat were included. The most dominant motif was (GGC/GCC)n, with the number of 64, following repeat motif were (AGC/GCT)n, (AG/CT) ?and (TGC/GCA)n.Primer sets wer designed for 15 ESTs sequences, for them , 14 wassourced from trinucleotid repeats, only 1 primer pair desigen from dinucleotide. Several primers which have been checked in Porphyra haitanensis have products, two primer pairs can discrimiante wild Porphyra haitanensis Cixi Ningbo, and cultivated strain of Xiangshan. The result show that it' s effective to transfer microsatellite primers sourced from Porphyra yezoensis ESTs to amplified in Porphyra haitanensis genomeIn conclusion, the single cell clone method shortened time greatly compared with traditional pure line establishment method, additional , company with the selective-breeding excellent strain through somatic cell, the trait slection and genetic stable can be made in step.Also it' s easier to check pure line with the help of molecular marker.