Gene Expression In The Regeneration Process Of The Algae Plume And Its Photosynthesis Research

The extruded protoplasm of Bryopsis hypnoides Lamouroux can aggregate spontaneously in seawater to form numerous new protoplasts, and part of them can develop into mature algal thalli, which phenomenon is inexisted for higher plants. This process is the restructure of the contents of the protoplasm from the dissociated cell fractions into orderly new cells, and is the de novo synthesis process of cell membranes and walls. This study was performed on different aspects of this aggregation process as follows:1. The preliminary studies on the gene and protein differential expressions of the protoplasts regeneration. Using realtime PCR, we quantified the expression level of the rbcL, psbO and lectin genes, results showed that the expression trends of those three genes were quite different during the protoplasts regeneration progress, presented many ups and downs phenomenon; However, mRNA expression of those three genes were all the lowest after 6 h onset of culture, this maybe related to the complex developmental stages during the protoplasts regeneration. The SDS-PAGE results showed that there were any changes on the distribution of the total soluble proteins after 6 h of cultivation. All the above results proved that the regulation of the protoplasts regeneration is multi-level, not only on the genomic level, but also on the protein levels.2. The sequencing of the B. hypnoides complete chloroplast genome. Chloroplasts from B. hypnoides were isolated and purified by differential centrifugation and sucrose density gradient centrifugation, and the chloroplast genome DNA (about 150kb) of B. hypnoides was extracted by high ion and low pH method combined with CsCl density gradient centrifugation. Then we constructed the genome library of the chloroplast genome DNA, sequenced the complete sequence of this cpDNA, and carried comparative genome and phylogenetic analyses on it.3. The photosynthetic characteristics about the protoplasts regeneration, the thalli and isolated protoplasts of B. hypnoides during dehydration. The results showed that the pigment contents, the ratios of photosynthesis rate to respiration rate, and the photosynthetic activity decreased gradually along with the regeneration progress, there was a relationship between the size of regenerated sub-protoplasts/protoplasts and their photosynthetic vigor, the middle-sized protoplasts which diametered 20-60μm had the higher photosynthetic activity and viability. During dehydration, the photosynthetic rates of B. hypnoides thalli were maintained before lost 30% of its initial water content, beyond which continued desiccation results in declined rates of both PSI and PSII. However, the isolated protoplasts showed a slight increase of Y (II) at 20% water loss, the rates decreased thereafter with declining water content. The isolated protoplasts retained some of the tolerance of the intact thalli, but their desiccation tolerance was not as good as the thalli, suggesting that cell wall may play an important role in desiccation tolerance.