末水条斑紫菜以组织捣碎机粉碎,在10 mmol/L磷酸盐缓冲液(p H 6.8)中反复冻融3次,收集上清液,通过膨化床疏水层析技术分离R-藻红蛋白(R-PE),剩余残渣用于硫酸多糖提取。使用膨化柱分离,粗提液中约17%的藻红蛋白可以被回收,DEAE-Sepharos...末水条斑紫菜以组织捣碎机粉碎,在10 mmol/L磷酸盐缓冲液(p H 6.8)中反复冻融3次,收集上清液,通过膨化床疏水层析技术分离R-藻红蛋白(R-PE),剩余残渣用于硫酸多糖提取。使用膨化柱分离,粗提液中约17%的藻红蛋白可以被回收,DEAE-Sepharose离子交换树脂纯化后,约12%的R-藻红蛋白得到回收,光谱纯度大于3.2,纯化得率为0.66 mg/g鲜紫菜。吸收光谱、荧光发射光谱以及凝胶电泳均显示该藻红蛋白属于典型三峰型R-藻红蛋白。利用藻红蛋白提取后的残渣,每克条斑紫菜可回收到30.5 mg的粗多糖,其中硫酸基含量为21.8%。实验证明,微波辅助抽提与热水浸提对硫酸多糖得率无显著影响。纯化的R-藻红蛋白与紫菜硫酸多糖可用于食品、化妆品添加,或药品、保健品的开发。本文对探索次等紫菜生物质的综合、高值化利用,进一步促进我国相关产品的开发与应用具有重要意义。展开更多
Pyropia haitanensis(T.J.Chang et B.F.Zheng) N.Kikuchi et M.Miyata( Porphyra haitanensis) is an economically important genus that is cultured widely in China.P.haitanensis is cultured on a larger scale than Pyropia yez...Pyropia haitanensis(T.J.Chang et B.F.Zheng) N.Kikuchi et M.Miyata( Porphyra haitanensis) is an economically important genus that is cultured widely in China.P.haitanensis is cultured on a larger scale than Pyropia yezoensis,making up an important part of the total production of cultivated Pyropia in China.However,the majority of molecular mechanisms underlying the physiological processes of P.haitanensis remain unknown.P.haitanensis could utilize inorganic carbon and the sporophytes of P.haitanensis might possess a PCK-type C 4-like carbon-fixation pathway.To identify micro RNAs and their probable roles in sporophyte and gametophyte development,we constructed and sequenced small RNA libraries from sporophytes and gametophytes of P.haitanensis.Five micro RNAs were identified that shared no sequence homology with known micro RNAs.Our results indicated that P.haitanensis might posses a complex s RNA processing system in which the novel micro RNAs act as important regulators of the development of different generations of P.haitanensis.展开更多
Pyropia haitanensis has prominent stress-resistance characteristics and is endemic to China. Studies into the stress responses in these algae could provide valuable information on the stress-response mechanisms in the...Pyropia haitanensis has prominent stress-resistance characteristics and is endemic to China. Studies into the stress responses in these algae could provide valuable information on the stress-response mechanisms in the intertidal Rhodophyta. Here, the effects of salinity and light intensity on the quantum yield of photosystem II in Py. haitanensis were investigated using pulse-amplitude-modulation fluorometry. Total RNA and genomic DNA of the samples under different stress conditions were isolated. By normalizing to the genomic DNA quantity, the RNA content in each sample was evaluated. The cDNA was synthesized and the expression levels of seven potential internal control genes were evaluated using qRT-PCR method. Then, we used geNorm, a common statistical algorithm, to analyze the qRT-PCR data of seven reference genes. Potential genes that may constantly be expressed under different conditions were selected, and these genes showed stable expression levels in samples under a salinity treatment, while tubulin, glyceraldehyde- 3-phosphate dehydrogenase and actin showed stability in samples stressed by strong light. Based on the results of the pulse amplitude-modulation fluorometry, an absolute quantification was performed to obtain gene copy numbers in certain stress-treated samples. The stably expressed genes as determined by the absolute quantification in certain samples conformed to the results of the geNorm screening. Based on the results of the software analysis and absolute quantification, we proposed that elongation factor 3 and 18S ribosomal RNA could be used as internal control genes when the Py. haitanensis blades were subjected to salinity stress, and that a-tubulin and 18S ribosomal RNA could be used as the internal control genes when the stress was from strong light. In general, our findings provide a convenient reference for the selection of internal control genes when designing experiments related to stress responses in Py. haitanensis.展开更多
基金Supported by the National High Technology Research and Development Program of China(863 Program)(No.2012AA10A406)the National Natural Science Foundation of China(Nos.41176134,41306151)the China Strategic Leading Special Science and Technology Academy(No.XDA11020404)
文摘Pyropia haitanensis(T.J.Chang et B.F.Zheng) N.Kikuchi et M.Miyata( Porphyra haitanensis) is an economically important genus that is cultured widely in China.P.haitanensis is cultured on a larger scale than Pyropia yezoensis,making up an important part of the total production of cultivated Pyropia in China.However,the majority of molecular mechanisms underlying the physiological processes of P.haitanensis remain unknown.P.haitanensis could utilize inorganic carbon and the sporophytes of P.haitanensis might possess a PCK-type C 4-like carbon-fixation pathway.To identify micro RNAs and their probable roles in sporophyte and gametophyte development,we constructed and sequenced small RNA libraries from sporophytes and gametophytes of P.haitanensis.Five micro RNAs were identified that shared no sequence homology with known micro RNAs.Our results indicated that P.haitanensis might posses a complex s RNA processing system in which the novel micro RNAs act as important regulators of the development of different generations of P.haitanensis.
基金Supported by the National Natural Science Foundation of China(Nos.41476140,41306151,41676157,41506172)the Strategic Leading Science and Technology Projects of Chinese Academy of Sciences(No.XDA11020404)+1 种基金the China Postdoctoral Science Foundation(No.2015M582153)the Science and Technology Plan of Jiangsu Province(No.BE2016330)
文摘Pyropia haitanensis has prominent stress-resistance characteristics and is endemic to China. Studies into the stress responses in these algae could provide valuable information on the stress-response mechanisms in the intertidal Rhodophyta. Here, the effects of salinity and light intensity on the quantum yield of photosystem II in Py. haitanensis were investigated using pulse-amplitude-modulation fluorometry. Total RNA and genomic DNA of the samples under different stress conditions were isolated. By normalizing to the genomic DNA quantity, the RNA content in each sample was evaluated. The cDNA was synthesized and the expression levels of seven potential internal control genes were evaluated using qRT-PCR method. Then, we used geNorm, a common statistical algorithm, to analyze the qRT-PCR data of seven reference genes. Potential genes that may constantly be expressed under different conditions were selected, and these genes showed stable expression levels in samples under a salinity treatment, while tubulin, glyceraldehyde- 3-phosphate dehydrogenase and actin showed stability in samples stressed by strong light. Based on the results of the pulse amplitude-modulation fluorometry, an absolute quantification was performed to obtain gene copy numbers in certain stress-treated samples. The stably expressed genes as determined by the absolute quantification in certain samples conformed to the results of the geNorm screening. Based on the results of the software analysis and absolute quantification, we proposed that elongation factor 3 and 18S ribosomal RNA could be used as internal control genes when the Py. haitanensis blades were subjected to salinity stress, and that a-tubulin and 18S ribosomal RNA could be used as the internal control genes when the stress was from strong light. In general, our findings provide a convenient reference for the selection of internal control genes when designing experiments related to stress responses in Py. haitanensis.
