RAPD技术在龙须菜遗传多样性研究中的应用 Ⅰ总DNA的提取及RAPD反应条件的优化

比较两种从龙须菜中提取总ＤＮＡ的方法,这两种方法得到的染色体ＤＮＡ具有较好的完整性,从ＤＮＡ的产量、蛋白质含量等指标可以看出,ＣＴＡＢ法优于蛋白酶Ｋ法。实验对龙须菜ＲＡＰＤ反应条件进行了优化：在２５μＬ反应体系中,含有Ｍｇ２＋２．５ｍｍｏｌ／Ｌ,ｄＮＴＰ１００μｍｏｌ／Ｌ,总ＤＮＡ２５ｎｇ,引物０．２μｍｏｌ／Ｌ和Ｔａｑ酶１Ｕ,经９４℃变性１ｍｉｎ,３５℃退火１ｍｉｎ,７２℃延伸２ｍｉｎ,３５个循环,得到稳定的ＲＡＰＤ扩增图谱。     

江蓠属藻胆体的研究 Ⅱ.藻胆体的荧光光谱特性

报道以青岛野生型龙须菜（ｗｔ）、黄色突变体（ｙｅ１００）、４株绿色突变体（ｇｒ１８０、ｇｒ１８４、ｇｒ１８６、ｇｒ１８８）、委内瑞拉野生型龙须菜（ｌｖ）、南非野生型龙须菜（ｓａ）及真江蓠（ｇａ）为材料,用蔗糖密度梯度离心法得到完整的藻胆体。用５４０ｎｍ激发,野生型的荧光发射峰位于５７８ｎｍ和６７０ｎｍ,相对荧光强度ＦＡＰＢ／ＦＰＥ的比值为１．５０。几种材料的ＰＥ和ＡＰＢ荧光发射峰没有明显区别（不超过２ｎｍ）,但两峰荧光强度比值显著差异。结果显示ｌｖ的ＦＡＰＢ／ＦＰＥ比值最低,只有１．２０,ｇｒ１８６的比值最高,为５．６７,野生型中ｓａ也比较高,为４．７９。用６７０ｎｍ激发,野生型龙须菜的荧光社发光谱有５个峰,分别为４９９ｎｍ、５４０ｎｍ、５６８ｎｍ、６１４ｎｍ、６５０ｎｍ,与其吸收光谱一一对应。野生型和突变体、不同产地龙须菜及真江蓠的荧光发射峰的波长都与其吸收光谱相对应。     

酞酸酯对龙须菜的生态毒理研究

研究了酞酸酯类化合物对龙须菜(Gracilaria lemaneiformis)生长的影响以及酞酸酯在龙须菜体内的积累状况。结果表明,在邻苯二甲酸二甲酯(DMP)、邻苯二甲酸二乙酯(DEP)、邻苯二甲酸二丁酯(DBP)、邻苯二甲酯二异辛酯(DEHP)4种酞酸酯混合质量浓度为0.05,0.10,0.20和0.40 mg/L的暴露条件下,龙须菜的相对生长速率和叶绿素a含量均受到了明显影响。与对照组比较,在高质量浓度处理组(0.1~0.4 mg/L)中,两指标分别下降了18.4%~21.3%和10.4%~15.3%,致毒效应与酞酸酯暴露浓度呈正相关。在受控水体的暴露试验中,利用气相色谱(GC-FID)分析表明,4种酞酸酯类(DMP,DEP,DBP和DEHP)在龙须菜体内均有一定程度的积累,其残留量分别为0~0.14 mg/kg,0~0.83 mg/kg,0.11~1.89 mg/kg,0.01~10.4 mg/kg,其积累程度与酞酸酯分子结构相关,侧链较长的酞酸酯比侧链较短的积累量要大,并随处理浓度的增加和时间推移呈增加趋势。     

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.     

Development of a SCAR marker for male gametophyte of Gracilariopsis lemaneiformis based on AFLP technique

The red alga Gracilariopsis lemaneiformis （Bory） is an economically valuable macroalgae. As a means to identify the sex of immature Gracilariopsis lemaneiformis, the amplified fragment length polymorphism （AFLP） technique was used to search for possible sex- or phase-related markers in male gametophytes, female gametophytes, and tetrasporophytes, respectively. Seven AFLP selective amplification primers were used in this study. The primer combination E-TG/M-CCA detected a specific band linked to male gametophytes. The DNA fragment was recovered and a 402-bp fragment was sequenced. However, no DNA sequence match was found in public databases. Sequence characterized amplified region （SCAR） primers were designed from the sequence to test the repeatability of the relationship to the sex, using 69 male gametophytes, 139 female gametophytes, and 47 tetrasporophytes. The test results demonstrate a good linkage and repeatability of the SCAR marker to sex. The SCAR primers developed in this study could reduce the time required for sex identification of Gracilariopsis lemaneiformis by four to six months. This can reduce both the time investment and number of specimens required in breeding experiments.     

龙须菜藻体匍匐组织的生理特性与功能研究

于１９９２年９月－１９９４年８月,在模拟潮间带生态条件的培养过程中,对龙须菜生长和发育的过程进行长期的连续性观察研究。研究中发现,以往被称为“固着器”的藻体匍匐组织具有特殊生理特性;它是孢子萌发后首先形成的部分,是直立藻体产生的基础;当夏末和深冬季节到来时,直立藻体通常成熟,放散孢子后逐渐衰退,这种趋势从藻体尖端开始逐渐向基部发展,最终保留匍匐组织度过逆境。待温度适宜时又从匍匐组织上萌生新的直立枝     

光强和盐度对龙须菜生长及光合特性的影响

持续降雨会引起光照强度和海水表层盐度的变化。近年来,受梅雨影响,福建省龙须菜(Gracilariopsis lemaneiformis)栽培产量波动明显。为揭示其原因,本实验设置3个盐度梯度(低盐：16,中盐：24,高盐：32)和两个光强水平[低光：30μmol/(m²·s),高光：120μmol/(m²·s)],共6个不同处理组合,研究其对龙须菜生长和光合特性的影响。结果发现：(1)盐度、光照强度和二者交互作用对龙须菜的相对生长速率和净光合速率均具有显著影响(P<0.05),适当降低盐度、增加光照有利于提高藻体相对生长速率和净光合速率,同时盐度适当降低条件下藻体呼吸速率被显著抑制;(2)光照强度和盐度对龙须菜有效光合量子产率、最大相对电子传递速率、光能利用效率均具有显著影响(P<0.05),在光照增加条件下有效光合量子产率和最大相对电子传递速率随盐度增加呈先增加后降低的趋势,光照强度和盐度的交互作用仅对光能利用效率影响不显著(P>0.05);(3)盐度对龙须菜的光合色素含量具有显著影响(P<0.05),在同一光强条件...     

氮素营养对龙须菜生长及色素组成的影响

在不同氮浓度及不同化合态氮条件下,探讨了龙须菜藻体的色素组成变化特性.结果为：不同化合态氮对龙须菜生长速率的影响差异不大,对藻体中藻红素含量的影响却差异显著;不同浓度氮（N）对龙须菜生长率的影响与所添加的氮浓度成正比,试验前期藻红素含量变化也是如此,但藻体中色素含量积累到一定浓度时就不再增加;另从藻红素与叶绿素a含量比值看,藻红素的增加主要是作为龙须菜体内氮贮存物质存在的.     

龙须菜配子体发育和不同品系四分孢子体四分孢子形成过程显微结构观察

龙须菜配子体和四分孢子体在生殖结构的形成和发育的过程中,出现一系列的细胞形态和结构层面的变化。本实验以龙须菜雌雄配子体和不同品系龙须菜四分孢子体为实验材料,采用连续徒手切片的方法,观察龙须菜配子体发育和不同品系四分孢子体四分孢子形成过程。观察到,雌雄配子体发育经过幼苗期、毛细胞期、性成熟期,幼苗期和毛细胞期雌雄配子体间无明显差异,进入性成熟期后,产生生殖结构时才能将其区分开。四分孢子起源于四分孢子体表皮层细胞,初始为鲜红色直径约5~10 μm的细胞,逐渐生长形成成熟的直径约20~25 μm的四分孢子囊,并分裂为4个四分孢子聚集在四分孢子囊中,在藻体外,四分孢子释放变形成球形,龙须菜四分孢子的形成和发育过程伴随着髓部细胞中红藻淀粉颗粒的减少。其中"981"龙须菜形成较多的畸形四分孢子,可能是其表现低育的一个原因。龙须菜自基部到尖端的表皮层细胞层数、髓部细胞数目和体积均发生连续的变化。细胞连接广泛存在于龙须菜表皮层、皮层和髓部细胞之间,但各层细胞连接大小、数目、长度等都存在差异。释放的四分孢子可能会附着在四分孢子体上,固着器覆盖包绕四分孢子体甚至侵入到破损有伤口的四分孢子体内,形成四分孢子体和配子体在同一株上的世代混杂现象,这也可能是导致龙须菜表现遗传复杂性的原因。     

龙须菜果孢子的紫外诱变及优势突变体的筛选

用紫外线辐射处理野生型龙须菜果孢子体放散刚附着的果孢子,并通过耐高温的筛选,初步获得了2株具有耐高温,生长快速等优良性状的四分孢子体突变株,命名为BZT-11和BZT-18,将其与981和野生型藻株进行了比较分析。结果显示,这2株突变体在实验室内和海上的日平均生长速率为均明显高于981和野生型藻株。在热胁迫时间内,BZT-11和BZT-18的丙二醛(MDA)含量均明显低于野生型,超氧化物歧化酶(SOD)活性明显高于野生型;而在整个胁迫过程中2株突变体与981藻株的MDA含量和SOD活性水平差异不显著。经过4h的热激后,藻株BZT-11和BZT-18的hsp70基因表达水平也显著高于野生型。这些结果进一步证明了该2株突变藻体的耐高温优势。本研究提供了耐高温突变体的筛选技术。