| Tissue culture for seed stock was studied on marine macroalgae Gracilariachouae. It is a kind of large red algae with significant economic and ecological value,and it has already been widely cultivated in Fujian coastal regions. However basicbiological research for G. chouae is still very limited at the moment. The seedlingincubation is hard to be conducted by spores and thus it mainly relies on vegetativepropagation for reproduction, which is not conducive to the expansion of farming areaand protection of germplasm resources. These problems seriously hampered thedevelopment of the cultivation of G. chouae. To supplement basic biological researchand seek for a more economical and efficient seedling incubation method, a series ofstudies were carried out for G. chouae cultured under laboratory conditions.Appropriate growth light intensity and temperature conditions were investigated.Besides tissue culture techniques were applied to explore the clone induction andseedling incubation method of G. chouae on several aspects, such as sterile processingof explants, optimization of physical and chemical conditions of tissue culture,induction, preservation and redifferentiation of filaments etc. The main researchmethods and results are as follows:1. A two-factorial experiment was designed to study the effects of temperature(10,15,20, and25℃) and light intensity (40,80,120,160and200μmol·m-2·s-1) onthe growth and biochemical composition of G. chouae. And the determination of therelative growth rate, photosynthetic pigment content, malondialdehyde (MDA) andsuperoxide dismutase (SOD) content and other indicators were carried out. The resultsshowed that temperature, light intensity, and their interactions had significant effectson the growth of G. chouae, and the effects of temperature was greater than those oflight intensity. A higher growth rate of G. chouae was observed at120μmol·m-2·s-1and20℃. Under such condition, G. chouae could reach to the maximum growth rateof0.038g·g-1·d-1. In addition, both temperature and light intensity had significanteffects on the chlorophyll-a and phycobiliprotein of G. chouae. In general, the photosynthetic pigment contents of G. chouae increased with the rise of temperature,while it increased initially and then dropped down with the rise of light intensity. Thecontents of phycoerythrin (PE), phycocyanin (PC), allophycocyanin (APC), andchlorophyll a were1.12mg/g,0.272mg/g,0.216mg/g and1.82mg/g separately whenthe light intensity was120μmol·m-2·s-1and temperature was25℃. Temperature, lightintensity, and their interactions had significant effects on the SOD and MDA contentsof G. chouae. And the effect of temperature on the content of MDA was greater thanthat of light intensity. The MDA content of G. chouae increased initially and thendropped down with increasing light intensity and temperature. The content of SODincreased with increasing temperature and light intensity. And light intensity was themain factor affecting the SOD content of G. chouae.2. The explants were cultured under different conditions, such as methods ofdetoxification, length, temperatures, light intensities, mediums and hormones. Theresults are as follows: a set of suitable explants detoxification methods for G. chouaehas been established. Firstly, pretreated explants were ultrasonic cleaned for15minutes, and then soaked in a1%povidone iodine solution for5min. After washingaway disinfectant reagent on the surface of explants, the explants were inoculated intothe f/2medium containing antibiotics for other24hours. This method can greatlyreduced the damage to explants which was caused by the detoxification andsignificantly increased the survival rate which is vital to obtain desired results in thefurther induction process. The most suitable explants length is10mm, in which sizethe explants had very ideal survival rate of100%and budding rate of98%. Theoptimum temperature and light intensity for inducing callus from G. chouae blockwere ranged from20℃to25℃and40~120μmol·m-2·s-1respectively. Under thiscondition, callus induction rate is higher than70%and budding rate higher than95%.Additionally, the induction of filaments and germination of propagules were affectedsignificantly when1-NAA was added in liquid medium.75%explants inductedfilaments, while no filaments were inducted in control groups.3. The filaments were inoculated in liquid medium and solid-liquid medium, andwere cultured under different temperatures and light intensities. The results showedthat the filaments in liquid medium maintained a rapid growth. The optimumtemperature is around20~25℃and the optimum light intensity is around80~120μmol·m-2·s-1. Maximum relative growth4rate reached0.10g.g-1. d-1. 4. Filaments’ fragments were cultured stationarily in PES liquid medium. Thecells of the filament’s fragment adherent to the substratum gave rise to crusts. Andgermination was obtained in40days through indoor culture. |
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