Isolation of Protoplasts from Undaria pinnatifida by Alginate Lyase Digestion

1　Introduction　Protoplastsserveasabasicandversatiletoolforge neticengineeringandbiochemicalresearchforseveralreasons :theymayregeneratetheirwallsandprovideamodelsystemforstudyingwallbiogenesis ,theirlysisprovidesagentlemeansofpreparingsubcellularor ganelles ,andthemembraneexpositionallowsgeneticmanipulationsinvolvingfusionoruptakeofnucleicacidstobepossible .Theseadvantagesarelessachiev ablewhenusingintactcells (Liciaetal.,1999) .Incomparisonwithlandplantsandunicellularalgae ,on lylimitedrepo…     

Studies on color type variants from mutagenized protoplasts ofPorphyra haitanensis Chang et Zheng &P. Yezoensis ueda (rhodophycease)

Isolated protoplasts from thalli ofPorphyra haitanensis andPorphyra yezoensis were treated with colchicine or irradiated by ultraviolet (UV). Several types of color variants were observed among the protoplast offspring. After treatment with colchicine: (1) 0.04–0.09% of red type variants inP. haitanensis were obtained; (2) The rate of red type variants and the variegated chimeral thalli composed of red type and wild type of sectors were 0.31–1.11% inP. yezoensis. After irradiation with UV: (1) 3.5–10.5% of red type variants inP. yezoensis were obtained: (2) 0.5–2.0% of red type variants and the variegated chimeral thalli composed of red type and wild type of sectors were obtained inP. haitanensis. Colchicine and UV’s mutangenic effects onP. yezoensis protoplasts were stronger than those onP. haitanensis protoplasts. The most efficient concentration of colchicine was 0.05%. The optimal length of UV-radiation was 1/2 min (radiation distance 5 cm). The red type variants induced by colchicine treatment grew faster than the wild type thalli. The clones of vegetative propagation from protoplasts of red type variants were still red type thalli. The red type variants will be good materials for genetic studies and improvement ofPorphyra strains.     

Genetic transformation of marine Actinomycete sp. isolate M048 and expression of a recombinant plasmid carrying the apc gene

1Introduction The ocean covering more than 70% of theearth’s surface represents a less explored environ-ment for microbial diversity(Yu etal., 2005). As agreat promising source for new natural productswhich have not been observed from terrestrial micro-o…     

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.     

海洋无脊椎动物消化酶的研究 Ⅰ、紫贻贝、日本蟳、滨螺消化酶的初步分析及其应用

本文用11种酶底物,测定了紫贻贝、日本蟳和滨螺消化系统的酶活性,并初步研究了这些消化酶的应用。实验表明:1.0％的紫贻贝消化酶在25℃、20分钟内可水解去除鲬鱼受精卵膜。1.0％的该消化酶加入等体0.5％的半纤维素酶和果胶酶,在30℃、60分钟内可制备出大量啤酒酵母原生质体。在同样条件下,1.5—2小时内,可去除大部分三角褐指藻的细胞壁。     

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.     

STUDIES ON COLOR TYPE VARIANTSFROM MUTAGENIZED PROTOPLASTS OFPORPHYRA HAITANENSIS CHANG ET ZHENG& P. YEZOENSIS UEDA (RHODOPHYCEASE )

Isolated protoplasts from thalli of Porphyra haitanensis and Porphyra yezoensis were treated with colchicine or irradiated by ultraviolet (UV ). Several types of color variants were observed among the protoplast offspring. After treatment with colchicine: (1) 0.04-0.09% of red type variants in P. haitanensis were obtained; (2) The rate of red type variants and the variegated chimeral thalli composed of red type and wild type of sectors were 6.31- 1.11% in P. yezoensis. After irradiation with UV: (1) 3.5- 10.5% of red type variants in P. yezoensis were obtained: (2) 0.5-2-0% of red type variants and the variegated chimeral thalli composed of red type and wild type of sectors were obtained in P. haitanensis. Colchicine and UV's mutangenic effects on P. yezoensis protoplasts were stronger than those on P. haitanensis protoplasts. The most efficient concentration of colchicine was 0.05%. The optimal length of UV-radiation was 1/2 min (radiation distance 5 cm). The red type variants induced, by colchicine tr     

长心卡帕藻原生质体分离、培养研究

In this study, protoplasts were successfully isolated from Kappaphycus alvarezii using snail enzymes, abalone enzymes and cellulase. The optimum enzymic ratio was fixed to be 20% of abalone enzyme, 12% of cellulase and the osmotic stabilizer was 2.0 mol/L glucose. The optimum enzymic hydrolysis conditions were found to be dark enzymolysis at 30°C continuing for 4.0 h. The resultant density and yield of protoplasts achieved 32.60×104 mL-1, 65.20×104 g-1 tissue for Kappaphycus alvarezii. Finally, under the temperature of 20°C, light intensity of 1 500–2 000 lx and photoperiod of 12 h/d, two developmental pathways were investigated:(1) callus-like cell mass and regenerated plantlet occurred on protoplast;(2) young shoots and calluslike cell mass occurred in tissue blocks after enzymolysis.     

微拟球藻原生质体制备与再生

综合考虑酶混合液处理时间和初始细胞密度两个因素,选取终浓度为4%半纤维素酶和2%崩溃酶的酶混合液对一株海洋经济微藻——微拟球藻(Nannochloropsis oculata)进行原生质体的制备与再生,Calcofluor White染料染色可在荧光显微镜下观察到完整的原生质体细胞。实验结果表明:同一初始密度藻细胞酶处理1～3 h制备率较高;酶处理相同时间较高初始密度(2×107～3×107cells/mL)的藻细胞制备率较高,并且原生质体在再生培养基上可再生,生长趋势与野生型细胞一致。考虑到酶处理时间过长或者密度过大会对原生质体的再生产生影响,本实验选择最适酶处理时间为1h,初始细胞密度为2×107cells/mL。     

坛紫菜和拟线形紫菜的原生质体融合及培养

应用PEG 法和电融合法进行了坛紫菜和拟线形紫菜的原生质体融合试验,探讨了PEG 的浓度、处理时间以及洗涤液种类与融合的关系。结果表明,PEG 法的融合率为4.9%～10.2%,电融合率高达32.3%。融合细胞经诱导长成多细胞团的愈伤组织,从愈伤组织上升出小紫菜。