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論文名稱: | Pseudomonas vesicularis MA103 菌株所產 Agarase PV-1b 純化與基因選殖至大腸桿菌表現之探討
| Studies on Purification of Pseudomonas vesicularis MA103 Agarase PV-1b and Its Gene Cloning and Expression in Escherichia coli
研究生: | 劉綉婷 Hsiu-Ting Liu
指導教授: | 潘崇良 Chorng-Liang Pan
學位類別: | 碩士
學校名稱: | 國立臺灣海洋大學
系所名稱: | 食品科學系
學號: | M95320017
畢業學年度: | 96
語文別: | 中文
關鍵字: | 純化 purification
| 基因選殖 gene cloning
| Agarase PV-1b Agarase PV-1b
| restriction-free cloning restriction-free cloning
| Escherichia coli 大腸桿菌
全文影像連結: | | (本論文20110729對校內公開)
| 論文全文(pdf)

論文頁數: | 98
摘要: | 本研究之目的在於探討 Pseudomonas vesicularis MA103 所產Agarase PV-1b 經純化及 N 端胺基酸定序，比對 MA103 genomic DNA 序列，以確認　Agarase PV-1b　的完整　DNA　序列，依序列設計專一性引子，以 restriction-free (RF) cloning 進行基因轉殖至大腸桿菌，轉殖表現後經親和性管柱初步純化重組 Agarase PV-1b (Agarase rPV-1b)。Agarase PV-1b 純化過程包括離心 (14,300 x g)、超過濾 (1 kDa MWCO)、陰離子交換層析、及膠體過濾層析，最終產物之純化倍率達 5.54 倍，分子質量約為 47 kDa。Agarase PV-1b 之 N 端胺基酸定序結果為 LDFNGYFRAGFGMNA。Agarase PV-1b 之生化特性測試結果顯示其水解 0.2% agarose 之最適作用溫度為 50oC，活化能為 21.76 J/mole，經酵素動力學分析得知，以 agarose 為基質時， Km 值為 9.7 mg/mL，KM 愈小表示酵素對基質之親和性愈高。Agarase PV-1b 對市售試藥 agar 及紅藻多醣具有基質特異性，水解洋菜糖 (Agarose) 及龍鬚菜、石花菜等藻類之熱萃多醣所得最終產物主要是新洋菜二糖和一個介於新洋菜四糖及六糖間的未知寡糖。根據 N 端胺基酸定序結果及蛋白質分子質量，自已完成定序的 P. vesicularis MA103 genomic DNA 中得到 Agarase PV-1b 核苷酸序列全長為 1299 bp，可轉譯出 432 個胺基酸。以 restriction-free (RF) cloning方法製得插入 Agarase rPV-1b 基因之重組載體 pET21b (+) 的 PCR 擴增產物，核苷酸序列全長約為 6000 bp，該重組載體經轉形反應 (Transformation) 殖入 E. coli BL21 (DE3) 後，已觀察到可表現agarase 活性之 Agarase rPV-1b (胞內粗酵素液)。轉形成功之轉形株 (Transformant) 經 isopropyl β-D-1-thiogalactopyranoside (IPTG, 1mM) 於 30oC 下誘導 6 hr 後，胞內粗酵素液可得最高酵素活性 (0.0012 U/mg protein)，該胞內粗酵素液經由親和性管柱層析處理後，得到初步純化之 Agarase rPV-1b (0.0760 U/mg protein)，此初步純化酵素液之 SDS-PAGE 結果顯示，於近似Agarase PV-1b 之分子質量 47 kDa 處可觀察到 Agarase rPV-1b。
| This study is aimed to purify Agarase PV-1b produced from Pseudomonas vesicularis MA103 and identified the N-terminal sequence of  Agarase PV-1b. According to the full genomic DNA sequencing result of P. vesicularis MA103 and N-terminal amino acid sequencing result of Agarase PV-1b, the full DNA sequence of Agarase PV-1b gene sequence was confirmed. Specific primers were designed to clone the Agarase PV-1b gene by restriction-free (RF) cloning, and the PV-1b gene was expressed in Escherichia coli system. The recombinant Agarase PV-1b (Agarase rPV-1b) was extracted from expression host to initially purify Agarase rPV-1b with affinity column.  Culture suspension of P. vesicularis MA103 was centrifuged (14,300 x g), ultrafiltration (1 kDa MWCO), and applied to anion-extrange chromatography and gel filtration chromatography to abtain Agarase PV-1b which was 5.54-fold purified and was analyzed to a molecular mass of 47 kDa.    The N-terminal amino acid sequence of Agarase PV-1b was determined to be LDFNGYFRAGFGMNA.  On the result of biochemical traits of Agarase PV-1b, the optimal temperature to degrade 0.2% agarose was 50oC, activation energy was calculated 21.76 J/mol, and Km was 0.97 mg/mL of agarose as substrate if Km value lower was regarded having lesser affinity to agarose.  Agarase PV-1b showed substrate specificity on red algal polysaccharides and agar products, and could degrade agarose and hot water extracted polysaccharides derived from Gracilaria sp. and Gelidium sp. to generate neoagarobiose and an unknowen oligosaccharide which had molecular size between neoagarotetraose and neoagarohexaose showed as the main end products.  According to N-terminal amino acid sequencing and molecular mass of Agarase PV-1b, the Agarase PV-1b gene was consisted of 1299 bp nucleotides encoding a protein of 432 amino acids from P. vesicularis MA103 full genomic DNA sequencing.  By restriction-free (RF) cloning method, it obtained the PCR amplification product of recombinant vector pET21b (+) inserting PV-1b gene about 6000 bp, and this plasmid was transformd into E. coli BL21 (DE3).  After isopropyl β-D-thiogalactopyranoside (IPTG, 1 mM) induction at 30oC for 6 hr, the intracellular crude enzyme of successful E. coli BL21 (DE3) transformant had the highest agarase activity (0.0012 U/mg protein, 1 unit (U) = 1 mg D-galactose equivalent/mL/min) and applied to affinity chromatography to obtain initially purified Agarase rPV-1b (0.0760 U/mg protein).  According to the result of SDS-PAGE, the molecular mass is 47 kDa which is close to Agarase rPV-1b.
論文目次: | 目錄    i
| 圖目錄    v
| 表目錄    vii
| 附錄目錄    viii
| 中文摘要    ix
| 英文摘要 (Abstract)    x
| 壹、前言    1
| 貳、文獻整理    3
| 一、洋菜酶 (Agarase)    3
| 1. 洋菜酶生產菌株及其分離來源    3
| 2. 分子質量    4
| 3. 洋菜酶轉殖    4
| 二、藻類多醣    5
| 1. 洋菜    5
| 2. 龍鬚菜    5
| 3. 海菜    6
| 4. 石花菜    6
| 三、藻類寡醣生理活性    7
| 1. 免疫性    7
| 2. 降低血脂    7
| 3. 益菌性    8
| 四、藻類寡醣組成成分鑑定    8
| 1. 薄層層析法 (Thin-layer chromatography, TLC)    8
| 2. 高效能液相層析法 (High-performance liquid
| chromatography, HPLC)    9
| 3. 磁核共振光譜法 (Nuclear magnetic resonance, NMR)    9
| 五、Restriction-free cloning    10
| 參、實驗設計    11
| 肆、實驗材料與方法    12
| 一、實驗材料    12
| 1. 原料    12
| 2. 實驗菌株與轉殖使用之載體    12
| 2.1 Pseudomonas vesicularis MA103菌株    12
| 2.2 蛋白質表現系統之生產菌株    12
| 2.3 載體來源    12
| 3. 藥品    13
| 4. 洋菜酶純化相關藥品配製    15
| 4.1 洋菜酶反應基質    15
| 4.2 DNS 溶液    15
| 4.3 膠過濾標準品    15
| 4.4 Wide molecular weight range-SDS (WMW-SDS) marker    16
| 4.5 SDS-polyacryamide gel electrophoresis
| (SDS-PAGE) 配製    16
| 4.6 1 kDa Ladder DNA marker    17
| 4.7 洋菜膠體電泳 (Agarose gel electrophoresis) 配製    17
| 5. 聚合酶鏈鎖反應試劑    18
| 6. 蛋白質純化套裝試劑    18
| 7. 培養基    18
| 8. 儀器設備    19
| 二、實驗方法    21
| 1.  Agarase PV-1 純化及定序    21
| 1.1 實驗菌株保存    21
| 1.2 實驗菌株活化    21
| 1.3 洋菜酶之生產及濃縮    21
| 1.4 洋菜酶的活性測定    22
| 1.5 蛋白質的定量    22
| 1.6 洋菜酶純化分離    22
| 1.6-(1) DE-52 陰離子交換樹脂層析    22
| 1.6-(2) SephacrylTM S-100 HR 膠體過濾層析    23
| 1.7 分子質量的分析    23
| 1.8 洋菜酶 N 端胺基酸定序    23
| 2. 洋菜酶生化性質測試    24
| 2.1 最適溫度    24
| 2.2 活化能    24
| 2.3 酵素動力學    24
| 2.4 基質特異性    24
| 3. 洋菜糖及藻類熱萃多醣之 Agarase PV-1b
| 水解物分析    25
| 3.1 藻類熱萃多醣之製備    25
| 3.2 洋菜糖及藻類熱萃多醣之 Agarase PV-1b
| 水解物之製備    25
| 3.3 薄層層析法 (Thin-layer chromatography, TLC)    25
| 3.4 高效能液相層析法 (High-performance liquid
| chromatography, HPLC)    25
| 4. 洋菜酶選殖及表現    26
| 4.1 菌株保存    26
| 4.2 菌株活化    26
| 4.3 勝任細胞之製備    26
| 4.4 載體 DNA 之製備    26
| 4.5 基因體 DNA 資料庫之建立    27
| 4.6 Agarase PV-1b 之基因序列        27
| 4.7 引子之設計    28
| 4.8 利用 restriction-free cloning (RF cloning) 擴增
| Agarase PV-1b 基因序列    28
| 4.9 重組載體以熱休克 (Heat shock) 法進行轉形
| (Transformation) 及基因定序    31
| 4.10 重組洋菜酶之純化及表現    32
| 伍、結果與討論    34
| 一、 Agarase PV-1b 純化及定序    34
| 二、 Agarase PV-1b 生化特性    35
| 1. 最適作用溫度    35
| 2. 活化能    35
| 3. 酵素動力學    36
| 4. 基質特異性    36
| 三、洋菜糖 (Agarose) 及藻類熱萃多醣之 Agarase PV-1b
| 水解物分析    37
| 1. 薄層層析法    37
| 2. 高效能液相層析法    38
| 四、Agarase PV-1b 選殖與重組 Agarase PV-1b 表現    39
| 1. Agarase PV-1b 基因序列及選殖    39
| 2. 重組 Agarase PV-1b 之純化及表現    40
| 陸、結論    42
| 柒、參考文獻    44
|
參考文獻: | 田勝朗。1990。作用於心臟血管系統之水產食品成分。食品工業。30 (3): 29-34。
| 吳欣蕙。2007。Pseudomonas vesicularis MA103 菌株所產 agarases PV-1 與 PV-2 之純化與性質探討。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 吳紹祺。1999。海洋細菌所產 agarases 生產條件與海藻多醣經 agarase 分解所得寡醣類組成之探討。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 吳紹祺。2005。藻類多醣洋菜酶水解物及其發酵產物之生理活性研究。國立臺灣海洋大學食品科學系博士學位論文，基隆，台灣。
| 袁景道。2001。海洋細菌 Pseudomonas vesicularis MA103 之 agarases 發酵槽生產條件探討以及海藻多醣與經酵素水解所得寡醣之應用。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 馬盈瑜。2003。龍鬚菜多醣及其水解物生理活性之探討。國立屏東科技大學食品科學系碩士學位論文，屏東，台灣。
| 康新楷。2007。Aeromonas salmonicida MAEF108 所產 Agarase AS-II 之純化與特性研究暨經 Agarase AS-II 水解所得藻類寡醣之生理活性探討。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 梁鳳鈺。1999。Pseudomonas vesicularis MA103 菌株所產洋菜酶的純化與性質。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 陳再發、紀美連、洗宜樂、薛月娥。1989。海菜化學組成、原料貯藏及加工之研究。臺水試澎所報彙集。8: 57-68。
| 陳君如。2006a。海菜酒之製備及其生理活性功能探討。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 陳姿元。2006b。Aeromonas salmonicida MAEF108 菌株所產洋菜酶的純化與性質之探討。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 黃韻笙。1997。狹葉紫菜 (Porphyra dentata) 中瓊膠脂萃取及其物化特性。國立臺灣海洋大學水產食品科學研究所碩士學位論文，基隆，台灣。
| 黃霨禎。2005。龍鬚菜寡醣水解物之製備及其抗氧化與抗致突變性之探討與對雙叉桿菌生長影響。國立臺灣海洋大學食品科學系碩士學位論文，基隆，台灣。
| 蔡憲華。1977。實用海藻類的化學組成之研究。臺灣省立海洋學院水產製造研究所，基隆，台灣。
| 顏嘉怡。2001。海洋嗜熱洋菜分解細菌與其洋菜酶之定性研究。東吳大學微生物學系碩士學位論文，台北，台灣。
| Allouch, J., M. Jam, W. Helbert, T. Barbeyron, B. Kloareg, B. Henrissat, and M. Czjzek.  2003.  The three-dimensional structures of two -agarases.  J. Biol. Chem.  278: 47171-47180.
| Allouch, J., W. Helbert, B. Henrissat, and M. Czjzek.  2004.  Parallel substrate binding sites in a β-agarase suggest a novel mode of action on double-helical agarose.  Structure  12: 623-632.
| Araki, T., M. Hayakawa, Z. Lu, S. Karita, and T. Morishita.  1998b.  Purification and characterization of agarases from a marine bacterium, Vibrio sp. PO-303.  J. Mar. Biotechnol.  6: 260-265.
| Bradford, M. M.  1976.  A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.  Biochemistry  72: 248-254.
| Cannell, R. J. P.  1989.  Algal biotechnology.  Appl. Biochem. Biotechnol.  20: 85-105.
| Chen, H. M., L. Zheng, and X. J. Yan.  2005.  The preparation and bioactivity research of agaro-oligosaccharides.  Food Technol. Biotechnol.  43: 29-36.
| Dong, J., S. Hashikawa, T. Konishi, Y. Tamaru, and T. Araki.  2006.  Cloning of the novel gene encoding -agarase C from a marine bacterium, Vibrio sp. strain PO-303, and characterization of the gene product.  Appl. Environ. Microbiol.  72: 6399-6401.
| Dong, T., Y. Tamaru, and T. Araki.  2007.  Molecular cloning, expression, and characterization of a -agarase gene, agaD, from marine bacterium, Vibrio sp. strain PO-303.  Biosci. Biotechnol. Biochem.  71: 38-46.
| Enzyme Nomenclature Database.  2008.  Agarase.  http://us.expasy.org/cgi-bin/enzyme-search-de  (June 27, 2008).
| Freile-Pelegrín, Y., D. R. Robledo, and G. García-Reina.  1995.  Seasonal agar yield and quality in Gelidium canariensis (Grunow) seoane-camba (Gelidiales, Rhodophyta) from Gran Canaria, Spain.  J. Appl. Phycol.  7: 141-144.
| Hassairi, I., R. B. Amar, M. Nonus, and B. B. Gupta.  2001.  Production and separation of α-agarase from Alteromonas agarlyticus strain GJ1B.  Bioresour. Technol.  79: 47-51.
| Hols, P., A. Baulard, D. Garmyn, B. Delplace, S. Hogan, and J. Delcour.  1992.  Isolation and characterization of genetic expression and secretion signals from Enterococcus faecalis through the use of broad-host-range α-amylase probe vectors.  Gene  118:21-30.
| Hosoda, A., M. Sakai, and S. Kanazawa.  2003.  Isolation and characterization of agar-degrading Paenibacillus spp. associated with the rhizosphere of Spinach.  Biosci. Biotechnol. Biochem.  67: 1048-1055.
| Ito K., and K. Hori.  1989.  Seaweed:  Chemical composition and potential food uses.  Food Rev. Int.  5: 101-104.
| Kang, N. Y., Y. L. Choi, Y. S. Cho, B. K. Kim, B. S. Jeon, J. Y. Cha, C. H. Kim, and Y. C. Lee.  2003.  Cloning, expression and characterization of a β-agarase gene from a marine bacterium, Pseudomonas sp. SK.38.  Biotechnol. Lett.  25: 1165-1170.
| Kazlowski B., Y. T. Ko, and C. L. Pan.  2007.  Comparison of two agaro-oligosaccharide separation and purification systems in the quantification of their oligomer products generated from agarose by HCl digestion.  Annual Meeting of Fisheries Society of Taiwan, Kaohsiung, Taiwan.  p. 151.
| Kim, B. J., H. J. Kim, S. D. Ha, S. H. Hwang, D. S. Byun, T. H. Lee, and J. Y. Kong.  1999.  Purification and characterization of β-agarase from marine bacterium Bacillus cereus ASK202.  Biotechnol. Lett.  21: 1011-1105.
| Kirimura, K., N. Masuda, Y. Iwasaki, H. Nakagawa, R. Kobayashi, and S. Usami.  1999.  Purification and characterization of a novel β-agarase from an alkalophilic bacterium, Alteromonas sp. E-1.    J. Biosci. Bioeng.  87: 436-441.
| Kobayashi, T., S. Ito, and K. Horikoshi.  2005.  Novel and useful enzymes from deep-sea and deep-subsurface microorganisms.  International Symposium on Extremophiles and Their Application 2005, Yokosuka, Japan.  pp. 58-65.
| Kong, J. Y. 2001. Production and functional properties of agarooligosaccharides.  Proceedings of Symposium on Scientific Study and Industrialization of Health Food, Health Food Society of Taiwan, Taipei, Taiwan.  pp. 60-81.
| Leon, O., L. Quintana, G. Peruzzp, and J. C. Slebe.  1992.  Purification and properties of an extracellular agarase from Alteromonas sp. strain C-1.  Appl. Environ. Microbiol.  58:4060-4063.
| Ma, C., X. Lu, C. Shi, J. Li, Y. Gu, Y. Ma, Y. Chu, F. Han, Q. Gong, and W. Yu.  2007.  Molecular cloning and characterization of a novel -agarase, AgaB, from marine Pseudoalteromonas sp. CY24.  J. Biol. Chem.  282: 3747-3754.
| Maeshige, S.  1962.  Chemical studies on the green alga, Monostroma nitidum wittrock-I.  Component sugars of the mucilage (part 1).  Bull. Jpn. Soc. Sci. Fish.  28: 326-334.
| Marino-Soriano, E.  2001.  Agar polysaccharides from Gracilaria species (Rhodophyta, Gracilariaceae).  J. Biotechnol.  89: 81-84.
| Mathews, C. K., K. E. Holde and K. G. Ahern.  Biochemistry.  1999.  3rd ed.  Addison Wesley Longman, SF, USA.  p.p. 360-381.
| Merck KGaA.  1998.  pET-21 a-d(+) Vector map.  http://www.merckbiosciences.com/docs/docs/PROT/TB036.pdf  (May 30, 2008).
| Morrice, L. M., M. W. McLean, F. B. Williamson, W. F. Long.  1983a.  β-agarase I and II from Pseudomonas atlantica. Purification and some properties.  Eur. J. Biochem.  135: 553-558.
| Morrice, L. M., M. W. McLean, W. F. Long, and F. B. Williamson.  1983b.  -agarases I and II from Psseudomonas atlantica. Substrate specificites.  Eur. J. Biochem.  137: 149-154.
| Norziah, M. H. and C. Y. Ching.  2000.  Nutritional composition of edible seaweed Gracilaria changgi.  Food Chem.  68:  69-76.
| Ohta, Y., Y. Nogi, M. Miyazaki, Z. Li, Y. Hayada, S. Ito, and K. Horikoshi.  2004.  Enzymatic properties and nucleotide and amino acid sequences of a thermostable β-agarase from the novel marine isolate, JAMB-A94.  Biosci. Biotechnol. Biochem.  68: 1073-1081.
| Oppert, B.  2006.  Two-dimensional analysis of proteinase activity.    J. Biochem. Biophys. Methods  67:173-179.
| Osumi, Y., M. Kawai, H. Amano, and H. Noda.  1997.  Isolation and indentification of a porphyran-decomposing bacterium.  Nippon Suisan Gakkaishi  63: 709-714.
| Osumi, Y., M. Kawai, H. Amano, and H, Noda. 1998. Effect of oligosaccharides from prophyran on in vitro digestions, utilizations by various intestinal bacteria, and levels of serum lipids in mice.  Nippon Suisan Gakkaishi  64: 98-104.
| Penaflorida, V. D. and N. V. Golez.  1996.  Use of seaweed meals from Kappaphycus alvarezii and Gracilaria heteroclada as binders in diets for juvenile shrimp Penaeus monodon.  Aquaculture  143:  393-401.
| Potin, P., C. Richard, C. Rochas, and B. Kloareg.  1993.  Purification and characterization of the -agarase from Alteromonas agarlyticus (Cataldi) comb. nov., strain GJ1B.  Eur. J. Biochem.  214:  599-607.
| Renn, D.  1997.  Biotechnology and the red seaweed polysaccharide industry: Status, needs and prospects.  Trends Biotechnol.  15: 9-14.
| Sambrook, J. and Russell D. W.  2006.  The Condensed Protocols from Molacular Cloning: A Laboratory Manual.  Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.
| Schroeder, D. C., M. A. Jaffer, and V. E. Coyne.  2003.  Investigation of the role of a β (1-4) agarase produced by Pseudoalteromonas gracilis B9 in eliciting disease symptoms in the red alga Gracilaria gracilis.  Microbiology  149: 2919-2929.
| Singh, T.  1992.  Agar and Agar Production.  INFOFISH, Kuala Lumpur, Malaysia  pp. 1-25.
| Stedman, T. L.  1982.  Stedman’s Medical Dictionary.  24th ed. Williams and Awaikins, London, UK.  p. 30.
| Stiles, M. E., and W. H. Holzapfel.  1997.  Lactic acid bacteria of foods and their currect taxonomy.  Int. J. Food Microbiol.  36: 1-29.
| Sugano, Y., I. Terada, M. Arita, M. Noma, and T. Matsumoto.  1993a.  Purification and characterization of a new agarase from a marine bacterium, Vibrio sp. strain JT0107.  Appl. Environ. Microbiol.  59: 1549-1554.
| Sugano, Y., T. Matsumoto, H. Kodama, and M. Noma.  1993b.  Cloning and sequencing of agaA, a unique agarase 0107 gene from a marine bacterium, Vibrio sp. strain JT0107.  Appl. Environ. Microbiol.  59: 3750-3756.
| Sugano, Y., T. Mastsumoto, H. Kodama, and M. Noma.  1994.  Purification and characterization of a novel enzyme, -neoagarooligosaccharide hydrolase (-NAOS hydrolase), from a marine bacterium, Vibrio sp. strain JT0107.  J. Bacteriol.  176: 6812-6818.
| Suzuki, H., Y. Sawai, T. Suzuki, and K. Kawai.  2003.  Purification and characterization of an extracellular β-agarase from Bacillus sp. MK03.  J. Biosci. Bioeng.  95: 328-334.
| Swartz, M. N. and N. Gordon.  1959.  Agarase from an agar-digesting bacterium.  J. Bacteriol.  77: 403-409.
| Usov, A. I.  1992.  Sulfated polysaccharides of the red seaweeds.  Food Hydrocoll.  6: 9-23.
| van den Ent, F. and J. Lowe.  2006.  RF cloning: A restriction-free method for inserting target genes into plasmids.  J. Biochem. Biophys. Methods  67: 67-74.
| van der Meulen, H. J., and W. Harder.  1975.  Production and characterization of the agarase of Cytophaga flevensis.  Antonie van Leeuwenhoek  41: 431-447.
| Vera, J., R. Alvarez, E. Murano, J. C. Slebe, and O. Leon.  1998.  Identification of a marine agarolytic Pseudoalteromonas isolate and characterization of its extracellular agarase.  Appl. Environ. Microbiol.  64: 4378-4383.
| Wang, Y. H., G. L. Yu, X. M. Wang, Z. H. Lv, X. Zhao, Z. H. Wu, and W. S. Ji.  2006.  Purification and characterization of alginate lyase from marine Vibrio sp. YWA.  Acta Biochim. Biophys. Sin.  38: 633–638.
| Wu, S. C., and C. L. Pan.  2004.  Preparation of algal-oligosaccharide mixtures by bacterial agarase and their antioxidative properties.  Fish. Sci.  70: 1164-1173.
| Ye, N., H. Wang, and G. Wang.  2006.  Formation and early development of tetraspores of Gracilaria lemaneiformis (Gracilaria, Gracilariaceae) under laboratory conditions.  Aquaculture  254:  219-226.
| Yoshizawa, Y., A. Ametani, J. Tsunehiro, K. Nomura, M. Itoh, F. Fukui, and S. Kaminogawa. 1995. Macrophage stimulation activity of the polysaccharide fraction of a marine alga (Porphyra yezoensis): Structure-function relationships and improved solubility.  Biosci. Biotechnol. Biochem.  59: 1933-1937.
| Yoshizawa, Y., J. Tsunehiro, K. Nomura, M. Itoh, F. Fukui, A. Ametani, and S. Kaminogawa.  l996.  In vivo macrophage-stimulation activity of the enzyme-degraded water-soluble polysaccharide fraction from a marine alga (Gracilaria verrucosa).  Biosci. Biotechnol. Biochem.  60: 1667-1671.
| Young, K. S., S. S. Battacharjee, and W. Yaphe.  1978.  Enzymic cleavage of the -linkages in agarose, to yield agaro-oligosaccharides.  Carbohydr. Res.  66: 207-212.
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2010 年 3 月 8 日 22:56