Biomass and carbon storage of Gracilariopsis lemaneiformis （Rhodophyta） in Zhanshan Bay,Qingdao, China

Marine macroalgae can absorb carbon and play an important role in carbon sequestration. As an important economic macroalga, Gracilariopsis lemaneiformis has the potential to significantly affect carbon absorption and storage in wave-sheltered intertidal reef systems. However, detailed knowledge on seasonal biomass changes and carbon storage of G. lemaneiformis is lacking, especially in many small and scattered ecosystems. Considering the influence of human activities on wild distribution of G. lemaneiformis, the understanding of seasonal dynamics of an economically important species in nature is necessary. In this study, we first investigated seasonal variations in biomass, coverage area, and carbon storage during low tide from August 2011 to July 2012 in Zhanshan Bay, Qingdao, China. Furthermore, we estimated the carbon storage potential of wild G. lemaneiformis using light use efficiency(LUE). The results show that the standing biomass and coverage area changed significantly with season. However, seasonal variations in carbon content and water content were not obvious, with an average content of 35.1% and 83.64%, respectively. Moreover, carbon storage in individual months varied between 0.67 and 47.03 g C/m 2, and the value of carbon storage was the highest in August and June and the lowest in February. In Zhanshan Bay, LUE of G. lemaneiformis was only 0.23%. If it is increased to the theoretical maximum(5%–6%), the carbon storage will have an increase of at least 21 times compared with the current, which suggested that carbon storage of wild G. lemaneiformis had a high enhancement potential. The study will help to assess a potential role of G. lemaneiformis in reducing atmospheric CO2.     

A Comparison of Different Gracilariopsis lemaneiformis （Rhodophyta） Parts in Biochemical Characteristics,Protoplast Formation and Regeneration

Gracilariopsis lemaneiformis is a commercially exploited alga.Its filaceous thallus can be divided into three parts,holdfast,middle segment and tip.The growth and branch forming trend and agar content of these three parts were analyzed,respectively,in this study.The results showed that the tip had the highest growth rate and branched most,although it was the last part with branch forming ability.The holdfast formed branches earliest but slowly.Holdfast had the highest agar content.We also assessed the difference in protoplast formation and regeneration among three parts.The middle segment displayed the shortest enzymolysis time and the highest protoplast yield; whereas the tip had the strongest vitality of protoplasts formation.Juvenile plants were only obtained from the protoplasts generated from the tip.These results suggested that the differentiation and function of G.lemaneiformis was different.     

Inhibition of photosynthesis in the microalga Chaetoceros curvisetus （Bacillariophyta） by macroalga Gracilaria lemaneiformis （Rhodophyta）

We investigated the effects of dried macroalga Gracilaria lemaneiform （Rhodophyta） on photosynthesis of the bloom-forming microalga Chaetoceros curvisetus. C. curvisetus was cultured with different amounts of dried G. lemaneiformis under controlled laboratory conditions. We measured the photosynthetic oxygen evolution rate and established the chlorophyll a fluorescence transient （OJIP） curve coupled with its specific parameters. We observed concentration-dependent and time-dependent relationships between dried G. lemaneiformis and inhibition of photosynthesis in C. curvisetus. Co-culture with dried G. lemaneiformis also resulted in a decrease in the light-saturated maximum photosynthetic oxygen evolution rate （P~ax） in C. curvisetus, and a decrease in the OJIP curve along with its specific parameters; the maximum photochemical efficiency of PSII （FJFm）, the amount of active PSII reaction centers per excited cross section at t=0 and t=--tFM （RC/CS0 and RC/CSm, respectively）, the absorption flux per excited cross section at t=0 （ABS/ CS0）, and the efficiency with which a trapped exciton moves an electron into the electron transport chain （~u0）. The dark respiration rate （Rd） increased in C. curvisetus co-cultured with dried G. lemaneiformis. The JIP-test and the oxygen evolution results indicated that dried G. lemaneiJbrmis decreased the number of active reaction centers, blocked the electron transport chain, and damaged the oxygen-evolving complex of C. curvisetus. This result indicated that dried fragments of G. lemaneiformis could effectively inhibit photosynthesis of C. curvisetus, and thus, could serve as a functional product to control and mitigate C. curvisetus blooms.     

Isolation and callus formation of Gracilariopsis bailiniae (Gracilariales,Rhodophyta) protoplasts

Journal of Oceanology and Limnology - This paper reports the first successful isolation of protoplasts from Gracilariopsis bailiniae and their callus formation. The base solution type,...     

Effects of temperature on photosynthetic performance and nitrate reductase activity in vivo assay in Gracilariopsis lemaneiformis (Rhodophyta)

Journal of Oceanology and Limnology - Gracilariopsis lemaneiformis is an economically-valued species and widely cultured in China at present. After being acclimated to different growth temperatures...     

Expression of the Phycoerythrin Gene of Gracilaria lemaneiformis (Rhodophyta) in E. coli and Evaluation of the Bioactivity of Recombinant PE

Phycoerythrin (PE) is one of the most important proteins involved in light capturing during photosynthesis in red algae. Its potential biological activities had gained wide concerns. In the present study, tumor cytotoxic and hydroxyl radical assay were preformed to detect the bioactivity of recombinant PE. Recombinant plasmids pGEX-PE and pBGL were transformed into E.coli BL21 to make two recombinant strains BEX (pGEX-PE) and BGL (pBGL). PE expressing in BEX (pGEX-PE) was validated by SDS-PAGE and Western blotting analysis. SDS-PAGE analysis indicated that the PE-GST fusion protein was mostly inclusion bod- ies. Specific expression of PE was confirmed by Western blotting analysis. The recombinant E. coli BEX (pGEX-PE) cells were col- lected and sonicated. The supernatants were reserved for the tumor cytotoxic experiments. The result of tumor cytotoxic assay indi- cated that the supernatants containing PE had the activity of inhibiting the growth of Hela cells and with the increase of protein con- centration, the inhibiting rate increased from 37.31% to 63.26%, which showed significant difference from the control. Hydroxyl radical scavenging effect was tested with supernatants of BEX (pGEX-PE) and BGL (pBGL) cell lysates treated with sonication and heating. For the sonication samples, the scavenging rates of the supernatants of BEX (pGEX-PE) and BGL (pBGL) cell lysates were significantly higher than the negative control BL21(pGEX-4T) (P<0.02), and the scavenging rates increased slowly following the increase of the protein content. For the heating samples, except for the 0.2 mg mL-1 BGL (pBGL) products, the scavenging effects of the supernatants of BEX (pGEX-PE) and BGL (pBGL) cell lysates were stronger than that of negative control BL21(pGEX-4T). However, the effect intensity was not positively correlated with the increase of the protein concentration. Though a partially de- creased hydroxyl radical scavenging activity was led by heating, the biological activity was still retained and conspicuous. This re- search showed that phycoerythrin protein expressing in E. coli has the potential medical and sanitarian value.     

Identification of a Putative Tetrasporophyte-Specific Gene in Gracilaria lemaneiformis(Gracilariales, Rhodophyte)

A putative tetrasporophyte-specific gene, designated as SSH466 (GenBank accession No. DQ019223), was one of the genes identified in this work using suppression subtractive hybridization (SSH) method in Gracilaria lemaneiformis. The full length of the gene was obtained using SMART RACE strategy. Sequence analysis revealed that the gene had 1 019 nucleotides, including an open reading frame of 498 nucleotides encoding 166 amino acid residues, 158 nucleotides of 5' untranslated region and 363 nucleo- tides of 3' non-coding region. Protein motif and secondary structure prediction showed that there existed a transmembrane domain with a unique β-sheet. Thus, SSH466 protein might be a cross-membrane protein. Sequence homology search in the public GenBank databases did not reveal any significant match with SSH466. Virtual Northern blot analysis confirmed that it was a tetrasporo- phyte-specific gene.     

Use of rapd for detecting and identifyingPorphyra (Bangiales, Rhodophyta)

Randomly amplified polymorphic DNA (RAPD) technique was applied to assess the genetic variations and phylogenetic relationships in 4 species ofPorphyra. The samples were collected from the coast of Canada, Vietnam, Zhoushan, Fujian and Qingdao in China. Amplifications with 20 primers were carried out under predetermined optimal reaction conditions (samples were first heated at 94°C for 5 min. and followed by 45 cycles of 1 min at 94°C, 1 min at 36°C, and 2 min at 72°C, then held at 72°C for 10 min). The amplified products were scored as present (1) or absent (0) for each DNA sample and an index of genetic similarity (F) was calculated by using Nei & Li’s matching coefficient method (1979). The value of (1−F) was used to quantify the genetic distances between species and construct a phylogenetic tree. The relationship indicated by the UPGMA and NJ cluster analysis on the values of the genetic distance is in good overall agreement with classical taxonomy. The obvious differences between natural and cultivated population ofP. haitanensi suggest that variation or hybridization with other species occurred during the culture. Contribution No. 100 from the Experimental Marine Biology Laboratory, IOCAS. Contribution No. 3289 from the Institute of Oceanology, Chinese Academy of Sciences. The research supported by the Foundation of the Chinese Academy of Sciences.     

Three new species of Batrachospermum Roth (Batrachosper-maceae,Rhodophyta) in China

1 INTRODUCTION The first published records of genus Batra-chospermum Roth in China are from Jao (1941), when eleven taxa were reported. Since then, Shi, Hu and Kumano (1993), Shi (1994a, 1994b), Hua and Shi (1996), Xie et al. (1999, 2003), Xie and Shi (2003) have described in more detail the species from China. To date, 23 taxa of Batrachospermum Roth in China have been reported, of which, B. yunnanense, B. nothocladoideum and B. transito-rium are new species of this genus, and are…     

Formation and early development of tetraspores of Polysiphonia urceolata (Rhodomelaceae, Rhodophyta)

Polysiphonia urceolata is one type of potential commercial red seaweeds used for breeding and cultivation, because of its significant biochemical and biomedical application. However, the information of breeding and seedling incubation for cultivation is limited, especially the early development. In this study, tetrasporohyte and gametophyte of P. urceolata were taken as the study materials in Huiquan Bay, Qingdao, China. The cleaned and sterilized tetrasporophytes and gametophytes were pre-cultured in sterilized seawater, then nurtured at 18℃, 25 μmol photons m-2 s-1 in 12:12 h (light:dark) photoperiod. Continuous observation under microscope showed that the early development consists of bipolar division stage and seedling stage. In the division stage, tetraspores germinate into bipolar sporelings that further differentiate into a colorless rhizoidal portion and a lightly pigmented upright shoot. The lightly pigmented rhizoidal cell develops to a rhizoid and the larger pigmented cell transforms to an erect axis. In the seedling stage, several quasi-protuberances appear on the erect axis and form juvenile seedlings. The results demonstrate the culture of P. urceolata from tetraspores under laboratory conditions.     

Formation and early development of tetraspores of Polysiphonia urceolata （Rhodomelaceae, Rhodophyta）

Polysiphonia urceolata is one type of potential commercial red seaweeds used for breeding and cultivation, because of its significant biochemical and biomedical application. However, the information of breeding and seedling incubation for cultivation is limited, especially the early development. In this study, tetrasporohyte and gametophyte of P. urceolata were taken as the study materials in Huiquan Bay, Qingdao, China. The cleaned and sterilized tetrasporophytes and gametophytes were pre-cultured in sterilized seawater, then nurtured at 18°C, 25 μmol photons m⁻² s⁻¹ in 12:12 h (light:dark) photoperiod. Continuous observation under microscope showed that the early development consists of bipolar division stage and seedling stage. In the division stage, tetraspores germinate into bipolar sporelings that further differentiate into a colorless rhizoidal portion and a lightly pigmented upright shoot. The lightly pigmented rhizoidal cell develops to a rhizoid and the larger pigmented cell transforms to an erect axis. In the seedling stage, several quasi-protuberances appear on the erect axis and form juvenile seedlings. The results demonstrate the culture of P. urceolata from tetraspores under laboratory conditions. Supported by National Key Technology Support Program, Development Program of China (No.2006AA09Z21), National Natural Science Foundation of China (No. 40618001 and N_CUHK438/06) and Shandong Agricultural Seed Stock Breeding Project     

Cloning and Analysis of Calmodulin Gene from Porphyra yezoensis Ueda （Bangiales, Rhodophyta）

In order to understand the mechanisms of signal transduction and anti-desiccation mechanisms of Porphyra yezoensis, cDNA and its genomic sequence of Calmodulin gene (CaM) was cloned by the technique of polymerase chain reaction (PCR) based on the analysis of P. yezoensis ESTs from dbEST database. The result shows that the full-length cDNA of CaM consists of 603 bps including an ORF encoding for 151 amino acids and a terminate codon UGA, while the length of genomic sequence is 1231 bps including 2 exons and 1 intron. The average GC content of the coding region is 58.77%, while the GC content of the third position of this gene is as high as 82.23%. Four Ca²⁺ binding sites (EF-hand) are found in this gene. The predicted molecular mass of the deduced peptide is 16688.72 Da and the pI is 4.222. By aligning with known CaM genes, the similarity of CaM gene sequence with homologous genes in Chlamydomonas incerta and Chlamydomonas reinhardtii is 72.7% and 72.2% respectively, and the similarity of the deduced amino acid sequence of CaM gene with homologous genes in C. incerta and C. reinhardtii are both 71.5%. This is the first report on CaM from a species of Rhodophyta.     

Cloning and Analysis of Calmodulin Gene from Porphyra yezoensis Ueda (Bangiales, Rhodophyta)

In order to understand the mechanisms of signal transduction and anti-desiccation mechanisms of Porphyra yezoensiss,cDNA and its genomic sequence of Calmodulin gene (CaM) was cloned by the technique of polymerase chain reaction (PCR) based on the analysis of P. yezoensis ESTs from dbEST database. The result shows that the full-length cDNA of CaM consists of 603 bps including an ORF encoding for 151 amino acids and a terminate codon UGA, while the length of genomic sequence is 1231 bps including 2 exous and 1 intron. The average GC content of the coding region is 58.77%, while the GC content of the third position of this gene is as high as 82.23%. Four Ca2+ binding sites (EF-hand) are found in this gene. The predicted molecular mass of the deduced peptide is 16688.72 Da and the pI is 4.222. By aligning with known CaM genes, the similarity of CaM gene sequence with homologous genes in Chlamydomonas incerta and Chlamydomonas reinhardtii is 72.7% and 72.2% respectively, and the similarity of the deduced amino acid sequence of CaM gene with homologous genes in C. incerta and C. reinhardtii are both 71.5%. This is the first report on CaM from a species of Rhodophyta.     

Comparison in copper accumulation and physiological responses of Gracilaria lemaneiformis and G. lichenoides (Rhodophyceae)

Heavy metal pollution has become a worldwide problem in aquaculture. We studied copper (Cu2+ ) accumulation and physiological responses of two red algae Gracilaria lemaneiformis and Gracilaria lichenoides from China under Cu2+ exposure of 0-500 μg/L in concentration. Compared with G. lemaneiformis, G. lichenoides was more capable in accumulating Cu2+ , specifically, more Cu2+ on extracellular side (cell wall) than on intracellular side (cytoplasm) and in cell organelles (especially chloroplast, cell nucleus, mitochondria, and ribosome). In addition, G. lichenoides contained more insoluble polysaccharide in cell wall, which might promote the extracellular Cu2+ -binding as an efficient barrier against metal toxicity. Conversely, G. lemaneiformis was more vulnerable than G. lichenoides to Cu 2+ toxin for decreases in growth, pigment (chlorophyll a, chlorophyll b, phycobiliprotein, and β-carotene) content, and photosynthetic activity. Moreover, more serious oxidative damages in G. lemaneiformis than in G. lichenoides, in accumulation of reactive oxidative species and malondialdehyde, and in electrolyte leakage, because of lower antioxidant enzyme (superoxide dismutase and glutathione reductase) activities. Therefore, G. lichenoides was less susceptible to Cu2+ stress than G. lemaneiformis.     

Effects of elevated CO2 on the photosynthesis and nitrate reductase activity of Pyropia haitanensis (Bangiales,Rhodophyta) grown at different nutrient levels

Pyropia haitanensis,a commercially important species,was cultured at two CO2 concentrations(390×10- 6 and 700×10-6(parts per million)) and at low and high nutrient levels,to explore the effect of elevated CO2 on the species under nutrient enrichment. Results show that in CO2-enriched thalli,relative growth rate(RGR) was enhanced under nutrient enrichment. Elevated CO2 decreased phycobiliprotein(PB) contents,but increased the contents of soluble carbohydrates. Nutrient enrichment increased the contents of chlorophyll a(Chl a) and PB,while soluble carbohydrate content decreased. CO2 enrichment enhanced the relative maximum electronic transport rate and light saturation point. In nutrient-enriched thalli the activity of nitrate reductase(NRA) increased under elevated CO2. An instantaneous pH change in seawater(from 8.1 to 9.6) resulted in reduction of NRA,and the thalli grown under both elevated CO2 and nutrient enrichment exhibited less pronounced reduction than in algae grown at the ambient CO2. The thermal optima of NRA under elevated CO2 and/or nutrient enrichment shifted to a lower temperature(10–15°C) compared to that in ambient conditions(20°C). We propose that accelerated photosynthesis could result in growth increment. Nassimilation remained high in acidified seawater and refl ected increased temperature sensitivity in response to elevated CO2 and eutrophication.     

New phenomenon in early development of sporelings in Gracilaria asiatica Chang et Xia （Gracilariaceae, Rhodophyta）

1 INTRODUCTION The red alga Gracilaria belongs to Gracilari- aceae, Gigartinales, Florideophyceae, and includes over 100 recorded species, wildly distributing in tropical and temperate regions (Oliveira and Plastino, 1994), and is useful for producing val…     

Effects of salinity, light and temperature on growth rates of two species of Gracilaria （Rhodophyta）

Effects of temperature, salinity and light intensity on growth rates of Gracilaria lichenoides and G. tenuistipitata var. liui Zhang et Xia were tested. Eight to ten levels of each factor were first tested separately. The best growth rate was obtained under the conditions of 32°C, 30 and 240 μmol/(m²·s) for G. lichenoides, and 24°C, 20 and 200 μmol/(m²·s) for G. tenuistipitata, respectively. Then a uniform design was used to evaluate the optimal combinations of the three factors. The best conditions for the highest daily specific growth rates (% increase in wet weight) are determined to be 31.30°C, 32.10, and 287.23 μmol/(m²·s) for G. lichenoides (16.26%/d), and 25.38°C, 21.10, and 229.07 μmol/(m²·s) for G. tenuistipitata (14.83%/d), respectively. Supported by the 908 Special Program (908-02-04-07), the National Basic Research Program of China (973 Program, No. 2006CB400608), and K. C. Wong Magna Fund in Ningbo University     

Effects of salinity, light and temperature on growth rates of two species of Gracilaria (Rhodophyta)

Effects of temperature, salinity and light intensity on growth rates of Gracilaria lichenoides and G tenuistipitata var. liui Zhang et Xia were tested. Eight to ten levels of each factor were first tested separately. The best growth rate was obtained under the conditions of 32～C, 30 and 240 μmol/(m2·s) for G. lichenoides, and 24℃, 20 and 200 μmol/(m2.s) for G. tenuistipitata, respectively. Then a uniform design was used to evaluate the optimal combinations of the three factors. The best conditions for the highest daily specific growth rates (% increase in wet weight) are determined to be 31.30℃, 32.10, and 287.23 μmol/(m2·s) for G. lichenoides (16.26%/d), and 25.38℃, 21.10, and 229.07 μmol/(m2·s) for G. tenuistipitata (14.83%/d), respectively.     

Development of organelle single nucleotide polymorphism(SNP) markers and their application for the identification of cytoplasmic inheritance patterns in Pyropia yezoensis(Bangiales,Rhodophyta)

The genus Pyropia contains several important cultivated species.Genetic research in nori species has mainly focused on the cell nucleus,with few studies on organelles(chloroplast and mitocho ndria).Due to the high copy numbers of organelles in cells,which influence the development and traits of algae,it is necessary to study their genetic mechanism.In this study,the marine red alga yezoensis,an important economic macroalga,was selected as the study object.To investigate organelle(chloroplast and mitochondria) inheritance in P yezoensis,the wild type RZ(maternal strain) was cro ssed with the red mutant HT(paternal strain) and 30 color-sectors from 11 F1 gametophytic blades were examined.The complete chloroplast and mitochondrial genomes of the red mutant(HT) were assembled for the first time.One reliable and stable single nucleotide polymorphism(SNP) loci filtrated by bioinformatics analysis was used as a molecular marker for chloroplast and mitochondrial DNA,respectively,in subsequent experiments.PCR amplification and sequence analysis showed that the haplotypes of color-sectors detected were consistent with those of the maternal parent,confirming that both chloroplast and mitochondrial genomes were inherited maternally in P.yezoensis.The inheritance pattern of organelles in P.yezoensis can be used to guide the hybridization and breeding of nori.Additionally,the organelle SNP markers developed in this study can be applied in subsequent genetic research.     

Genetic improvement on Chinese shrimp （Fenneropenaeus chinensis）： growth and viability performance in F1 hybrids of different populations

Fenneropenaeus chinensis distributed in the Yellow Sea and Bohai Sea of China and the west coast of the Korean Peninsula. Different geographical populations represent potentially different genetic resources. To learn further the characteristics of different geographical population, crosses among two wild and three farmed populations were produced. The two wild populations were from the Yellow Sea and Bohai Sea (WYP), and the west coast of the Korean Peninsula and coast (WKN). The three farmed populations included the offspring of first generation of wild shrimp from coast in Korea (FKN), the Huang Hai (the Yellow Sea in Chinese) No. 1 (HH 1), and JK98. The phenotypes growth and survival rates of these populations were compared to confirm the feasibility for crossbreeding. The body length (BL), carapace length (CL), carapace width (CW), height of the second and third abdominal segment (HST), width of the second and third abdominal segment (WST), length of the first abdominal segment (LF), length of the last abdominal segment (LL), live body weight (BW), and survival rate were measured. Different combinations were statistically performed with ANOVA and Duncan's Multiple significant difference (P<0.05) in BL, CL, HST, LL, and BW; and insignificant difference (P0.05) in other growth traits and survival rate. The results of Duncan's Multiple Range Test are that BL and CL of was the best combination in all growth traits. Therefore, hybridization can introduce the variation to base populations. The systematic selection program based on additive genetic performance may be more effective than crossbreeding.