Cloning And Application Of The Nuclear Ribosomal DNA Cistron(nrDNA) Sequence Of Two Species Of Pyropia In China

Bangiaceae consist of two genus: Bangia and Pyropia, they are worldwide distribution and both of them are economically important seaweed. In this article, we chose Pyropia yezoensis and Pyropia haitanensis as the materials, and the complete nuclear ribosomal DNA cistron sequence of Pyropia haitanensis is cloned and analyzed. The nr DNA variation is compared for the nine cultivated variaties of Pyropia yezoensis in China. In addition, we made a similarity analysis of different parts of a piece of Pyropia yezoensis which collected from Rizhao, Shandong province, the result will provide an molecular evidence to the meiosis of Pyropia occurs during the germination of conchospore.The ribosomal RNA gene is a multi-gene family which is composed of SSU r DNA, non-coding regions including ITS1, 5.8S r DNA, ITS2, LSU r DNA and intergenic spacer(IGS). The gene cloning and sequencing methods were used to analyze the r DNA of Pyropia haitanensis which is cultivated in Putian, Zhejiang province and the total sequence is 15528 bp, through amplying c DNA, we found that the sequence of SSU r DNA is 2953 bp which including upstream intron(561bp) and downstream intron(555bp). The total sequence of LSU r DNA is 4444 bp including upstream intron(372bp) and downstream intron(1052bp), and the 5.8S r DNA gene is158 bp, the ITS1 is 331 bp and the ITS2 is 673 bp, the ITS of different individuals are almost identical. The structure of IGS is complex, so we designed six pairs of primers to amply the total sequence, and obtained the complete IGS(6969bp) through manual stitching.The IGS variation was compared for the nine main cultivated varieties of Pyropia yezoensis in China. The nine cultivars are ST No.1, ST No.2, ST No.3, ST No.4, DLNo.1, DL No.2, QD wild type, Y-H001 and Y-H002. The sequence analysis of four partial IGS amplicons ranged from 3628 bp to 3776bp; and the G+C content is 52.5%.Through similarity analysis and multiple sequence alignment of these partial IGS amplicons from the nine varieties, we have found 278 variable sites which account for approximately 7.5% of the whole IGS length. Three strains, ST No.1, ST No.2, DL No.1 were significantly different from the other strains. In addition, 56 positions experienced transitions or transversions. What’s more, we divided a piece of Pyropia yezoensis into ten parts, and used the partial IGS(834bp) to sequence and analyze.Through similarity analysis of the sequences of reproduction, nutrition, and base, we found that the thalli are either homozygotes or heterozygotes. The results indicate that Pyropia yezoensis is a chimera which consists of different genetic cells, the conclusion will provide an evidence that meiosis of Pyropia occurs during the germination of the conchospore.