【作者】 戴耀虹；

【导师】 边银丙；

【作者基本信息】 华中农业大学 ， 微生物学， 2011， 硕士

【摘要】 本试验以香菇(Lentinula edodes)4个栽培菌株[武香一号(W1)、L205、秋6、L271]和2个野生菌株(SHL12、SHL44)为试验材料,根据优势互补的原则形成4对杂交组合,采用单-单及单-双两种方式进行杂交,依据吃料速度和漆酶活性测定,筛选获得87个杂交子进行出菇试验,运用灰色关联度分析方法对供试菌种8个主要性状值[菌盖直径、菌盖厚度、菌柄长度、菌柄直径、平均单菇鲜重、单袋平均鲜菇产量、单袋平均菇数、出菇期]进行综合评价。从44个正常出菇的杂交菌株中,初步筛选出综合性状较优良的6个杂交菌株(XZ-3、XZ-8、XZ-40、XZ-74、XZ-81和XZ-87)进入中试。运用IRAP标记对杂交组合W1×SHL12的39个杂交子进行了遗传分析。采用扩增重复性好、谱带清晰的12对引物,扩增共获得128条带,其中127个多态性条带,多态性比例为99.2%。根据引物扩增图谱,采用软件NTSYS2.10e进行聚类分析并计算菌株间的遗传相似系数。结果显示,所有供试菌株在0.69的水平上聚为两类。第Ⅰ类菌株中,除XZ-27、XZ-28之外,均为单-单杂交获得的杂交子；第Ⅱ类菌株中,除XZ-5、XZ-23之外,均为单-双杂交获得的杂交子。菌株XZ-10与XZ-11出菇很少,个体较大,在0.88相似水平上聚为一类。XZ-3、XZ-8出菇数量适中,个大肉厚,产量远高于两亲本,与XZ-2在0.77水平上聚为一类。XZ-19、XZ-20、XZ-27出菇较少,个体较小,菌袋易吐黄水,在0.83水平上聚为一类。XZ-5、XZ-23两个菌株均未出菇,在0.83水平上聚为一类。XZ-7与XZ-17的菌丝均能长满菌袋,但栽培袋不起瘤状物,亦不转色,在0.86水平上聚为一类。试验结果表明,IRAP分析能较好地反映香菇种质的遗传多样性,依据IRAP指纹图谱进行的聚类分析,可用于香菇优良杂交菌株的初步筛选中,从而进一步提高杂交育种工作的效率。采用TRAP、ISSR、SSR、RAPD、SRAP分子标记的方法,对杂交组合中6个亲本菌株(武香一号、L205、秋6、L271、SHL12、SHL44)和55个单核体亲本(武香一号单核体16个、L205单核体7个、秋6单核体3个、L271单核体10个、SHL12单核体9个、SHL44单核体10个)进行遗传分析。28对扩增重复性好、谱带清晰的引物,在W1×SHL12组合中共扩增出912个条带,多态性条带数为850个,占总条带数的93.2%；在L205×SHLl2组合中共扩增出924条带,多态性条带数为877条,占总条带数的94.9%；在L205×SHL44组合中共扩增出863条带,多态性条带数为806条,占总条带数的93.4%；在秋6×L271组合中共扩增出780条带,多态性条带数为700条,占总条带数的89.7%。聚类结果很好的反映了各菌株间的亲缘关系,但亲本间的遗传距离与栽培试验中正常出菇的44个杂交子主要农艺性状值及灰色关联度法分析结果间的相关性均较低。用分子标记估算亲本遗传距离与杂种后代综合性状间的相关性存在诸多不确定性,不仅受遗传背景的影响,在一定程度上也受环境因子的影响。更多还原

【Abstract】 In this study, according to the principle of heterosis, test-strains of four cultivation strains W1, L205, Q6,271 and 2 wild strains SHL12, SHL44 were selected to form four pairs of complementary crossing combination, hybridized by the technique of Mon-Mon crossing and Di-Mon crossing breeding. Then according to the speed of Mycelial growth and laccase activity,87 hybrids were Selected from them. The method of grey correlation degree analysis was applicated to comprehensively analysis the 8 main agronomic characters of test-strains, which were pileus diameter, pileus thinkness, stipe length, stipe diameter, average fresh weight of a single fruitbody, average fresh yield of fruit body per bag, average numbers of fruitbody per bag and fruiting period. Six more excellent strains (XZ-3, XZ-8, XZ-40, XZ-74, XZ-81, XZ-87) selected from 44 normal fruiting of hybrid strains were further studied in the middle test.IRAP marker was applied to analyze the genetic diversity of 39 hybrid strains from the W1×SHL12 cross combination.12 pairs of IRAP primers were selected and a total of 128 fragments were attained, among which 127 fragments (rate of 99.2%) were polymorphic. Based on amplification map, software NTSYS2.10e was used to calculate the genetic similarity and analyze cluster results between the tested strains. The results show that, all the tested strains were clusted into 2 groups at 0.69 similarity level. the strains of all Mon-Mon crossings except XZ-27, XZ-28 were put into the first group, and all Di-Mon crossings except XZ-5, XZ-23 into the second group. XZ-10 and XZ-11 had few but big fruitbody, so they could be put into one group at 0.88 similarity level. XZ-3 and XZ-8 had big fruitbody, higher yield than their parents, clusted into one group with XZ-2 at 0.77 similarity level. XZ-19, XZ-20, XZ-27 had few and small fruitbody, cultivation bags of which were apt to spit out yellow water, classified into one group at 0.83 similarity level. XZ-5, XZ-23 have no fruitbody, classified into one group at 0.83 similarity level. XZ-7, XZ-17 covered with mycelium in bags completely, which had no tumor and no turn color, were clusted into one group at 0.86 similarity level. The results of IRAP analysis could well reflect the genetic diversity of the Lentinula edodes, and could guid for initial screening excellen hybrid strains of L. edodes strains, which could further improve the efficiency of hybrid breeding.We chosed 6 dicaryotic parents (W1, L205, Q6,271, SHL12, SHL44) and 55 monocaryotic parents (16 monocaryons of W1,7 monocaryons of L205,3 monocaryons of Q6,10 monocaryons of L271,9 monocaryons of SHL12,10 monocaryons of SHL44) as tested strains to analyze their genetic similarity by DNA Fingerprint with TRAP, ISSR, SSR, RAPD, SRAP.28 pairs of primers were selected to amplify the hybrid strains, and a total of 912 fragments were attained from the pair of W1×SHL12, among which 850 fragments were polymorphic, with the rate of 93.2%; 924 fragments from the pair of L205×SHL12, among which 877 fragments were polymorphic, with the rate of 94.9%; 863 fragments from the pair of L205×SHL44, among which 806 fragments were polymorphic, with the rate of 93.4%; 780 fragments from the pair of Q6×L271, among which 700 fragments were polymorphic, with the rate of 89.7%; According to amplification results, amplification products supported the genetic relationship among strains. However, genetic distance and the value of the main agronomic traits and sort results of grey relation degree were not significantly correlated. The method using molecular markers to estimate correlation of genetic distance and traits of hybrids exists many uncertainties, the results of which not only are affected by the genetic background, but also by environmental factors to some extent.更多还原