【作者】 张青；

【导师】 史全良；

【作者基本信息】 苏州大学 ， 植物学， 2011， 硕士

【摘要】 地衣是由真菌和藻类共生联合而成的有稳定形态和特殊结构的复合体,是目前生物界中最突出、最成功的共生现象的典范,被称为“生物的开路先锋”。目前梅衣科地衣中还未见有报道存在共生蓝藻,而且藻菌共生与生长基质的关系也未见报道。因此本研究采用形态学方法、化学显色法与分子生物学方法相结合鉴定了采自云南省的10个地衣样品,并对其共生藻进行了分离纯化培养及鉴定;探讨了藻菌共生组合与其生长基质之间的关系。采用GADPH和ITS基因序列构建系统发育树分析10个地衣样品(实验室编号分别为D1、D2、D3、D4、D5、D6、D7、D8、D9、D10。其中D1、D4、D5、D6、D9为树生地衣,D2、D10为土生地衣,D3、D7、D8为石生地衣)。结果表明,10个地衣样品隶属于茶渍目(Lecanorales)梅衣科(Parmeliaceae)大叶梅属(Parmotrema),D1、D3、D4、D5、D6、D7、D8、D9与大叶梅(Parmotrema tinctorum)亲缘关系最近,其中D3、D4、D6隶属于大叶梅(Parmotrema tinctorum),D2、D10隶属于大叶梅属(Parmotrema)的同一个种Parmotrema perlatum。分离纯化10个地衣共生藻(实验室编号为Z1、Z2、Z3、Z4、Z5、Z6、Z7、Z8、Z9、Z10),对其进行形态鉴定及rbcL、ITS基因序列分析构建系统发育树。结果表明,5个树生地衣共生藻样品Z1、Z4、Z5、Z6、Z9为念珠藻目(Nostocales)念珠藻科(Nostocaceae)念珠藻属(Nostoc);2个土生地衣共生藻样品Z2、Z10为四孢藻目(Tetrasporales)胶球藻科(Coccomyxaceae)胶球藻属(Coccomyxa);3个石生地衣分离藻样品Z3、Z7、Z8是绿球藻目(Chlorococcales)栅藻科(Scenedesmaceae)链带藻属(Desmodesmus)。另外,尽管Z1、Z5、Z9分别是从采自不同区域(姚安县及南华县)的地衣中分离纯化的,但亲缘关系最近;同样Z4、Z6亲缘关系最近;Z2、Z10亲缘关系最近,Z7、Z8亲缘关系最近。所以地衣共生菌与共生藻的共生关系与其生存的地理区域关系不大,但却和其生长基质具有明显的相关性。许多学者曾试图实现地衣菌藻共生的人工重建,然而,除了Stahl、Ahmadjian等人的实验之外,几乎没有成功的报道。Ahmadjian及Heikkila曾重复了Stahl关于壳状地衣(Crustaceous lichen)石果衣(Endocarpon pusillum)的人工重建实验,从而进一步证实了由该地衣的无藻子囊孢子和无菌共生藻细胞重建了共生联合之后能够形成具有子囊壳的地衣体。Stocker-W(o|¨)rg(o|¨)tter等在实验室土壤基质中成功再合成蓝藻型地卷属地衣叶状菌体。只有存在足够数量的共生光合生物细胞才能在琼脂平板中形成地衣真菌菌丝体。地衣的重建在土壤基物上较为成功。本研究结果表明,地衣共生藻类型与地衣生长基质具有明显的相关性,而与生存的地理区域关系不大,因此可以推测地衣菌藻人工重建困难可能是因为人工重建条件与地衣生长的基质和实际环境条件不同所致。地衣中目前报道的共生蓝藻主要存在于地卷目(Peltigerales),如地卷科(Peltigeraceae)、肺衣科(Lobarlaceae)、胶衣科(Collemataceae)、猫耳衣科(Leptogiummenziesii)等,梅衣科(Parrneliaceae)地衣中还未见有报道。本研究显示树生梅衣科地衣中存在共生念珠藻(Nostoc),梅衣科地衣共生藻为绿藻的传统观点有待完善。更多还原

【Abstract】 The lichen association is a mutualistic interaction between a mycobiont (fungal partner) and a phycobiont (cyanobacteria or green algal); the most successful example of symbiosis is known as the "pioneer organisms". Now there is cyanobacteria in Parmeliaceae which is not reported recently, furthermore, the relationship between algae-fungus symbiosis and growth matrix also has no reports. Ten specimens of lichens that collected from Yunnan province of China were identified by morphological identification, chemical development process and molecular biology method. Meanwhile, we isolated, purified and identified the phycobionts of the ten specimens. Furthermore, we discussed the relationships of phycobionts- mycobionts symbiosis on different growth matrixs.GADPH and ITS sequences were analyzed (The serial numbers of ten lichens specimens are respectively D1, D2, D3, D4, D5, D6, D7, D8, D9, D10 in the laboratory. D1, D4, D5, D6, D9 are corticolous lichens, D2, D10 are tecolous lichens, D3, D7, D8 are saxicolous lichens). The results show that the specimens of lichens are identified as Lecanorales, Parmeliaceae, Parmotrema. Thereinto, D1, D3, D4, D5, D6, D7, D8, D9 are closest affiliation with Parmotrema tinctorum, D3, D4, D6 are belonged to Parmotrema tinctorum, D2, D10 are belonged to Parmotrema perlatum.The phycobionts were isolated and purified (The serial numbers of ten phycobionts are Z1, Z2, Z3, Z4, Z5, Z6, Z7, Z8, Z9, Z10 in the laboratory) from ten specimens of lichens. Morphological identification and ITS, rbcL sequences were analyzed. The results show that five specimens of phycobionts from corticolous lichens, numbered Z1, Z4, Z5, Z6, Z9 respectively, are identified as Nostocales Nostocaceae Nostoc. Two specimens of phycobionts from tecolous lichens, Z2, Z10 are identified as Tetrasporales, Coccomyxaceae coccomyxa. Three specimens of phycobionts from saxicolous lichens, Z3, Z7, Z8 are identified as Chlorococcales Scenedesmaceae desmodesmus. Furthermore, the localities of lichens and phylogenetic analysis reveal that Z1, Z5, Z9 have closest affiliation. Similarly, it is closest affiliation that Z4, Z6 and Z2, Z10 and Z7, Z8. In concluding, mycobionts and phycobionts have rare relationships to the geographical origin, but it has obvious correlation with the growth matrix.Many scholars had tried to realize the artificial resynthesis of lichens. However, few successful experiments were reported besides Stahl and Ahmadjian. Ahmadjian and Heikkila repeated the Stahl’s experiment about the artificial resynthesis of Endocarpon pusillum, it further confirmed that no-algae ascospores and sterile phycobionts were able to form the thallus of lichens which had perithecium. Stocker-W(o|¨)rg(o|¨)tter researched that the resynthesis of foliose thalli of the cyanobacterial lichen Peltigera praetextata was achieved on a soil substratum under laboratory conditions. On agar plates mycelia formation by this lichen fungus only occurred when a sufficient number of photobiont cells was present. The artificial resynthesis of lichens is very successful on a soil substratum under laboratory conditions. Our results show that mycobionts and phycobionts have rare relationships to the geographical origin, but it has obvious correlation with the growth matrix. Therefore, the artificial resynthesis of lichens is very difficult. We consider that the artificial resynthesis conditions are different with growth matrix and actual environment.So far, according to reports, the cyanobacterias existed in Peltigerales only, such as Peltigeraceae, Lobarlaceae, Collemataceae, Leptogiummenziesii. There was no cyanobacteria in Parrneliaceae. However, this results show that the phycobiont is Nostoc in Parrneliaceae (corticolous lichens), Which improve the traditional view that all the phycobionts are green algae in Parrneliaceae.更多还原