Biodiversity Analysis Of Agrocybe Salicacola And Molecular Markers Selection For The Mutant Strains

Agrocybe salicacola of wild resources and non-sporulating mutant strain were studied,mainly including the genetic diversity,classification,identification of similar species,non-sporulating traits related to molecular marker development,sporeless mutant strain cytological,and morphological characteristics of fronts.Eighteen isolates of A.salicacola and A.cylindracea wild resources collected in Yunnan province as study materials,were analyzed in genetic diversity and relationship among species by use of AFLP.By using pheromone receptor,the diversity of A.salicacola and A.cylindracea were studied.The results are summaried as follows:● The genetic similarity coefficient of this kind of A.salicacola are greater than 0.87,the genetic relationships among isolates of relatively close,suggesting that genetic consistency of the high,a comparatively narrow genetic base,which indicates that the A.salicacola intraspecific differentiation is not obvious;● A.cylindracea genetic similarity coefficient ranges from 0.54 to 0.84,showing large genetic diversity among the A.cylindracea germplasm resources,which indicates that the differentiation within A.cylindracea is larger,which perhaps is resulted from geographical isolation or other factors.The genetic diversity of A.cylindracea for selection breeding provides a diverse genetic material and application prospect.● According to the results of phylogenetic tree of pheromone receptor nucleic acid sequences,the pheromone receptor nucleic acid information exists differences in the distribution of species.The information can be used as indicators of the species distinction.Based on the distribution of A.cylindracea and A.salicacola pheromone receptor,6 fragments amplified from A.salicacola YAASM0894 strain was located inⅠb,Ⅰc,Ⅰe and Ⅱb group.This phenomenon shows that the mating type gene information of A.salicacola YAASM0894 strain has more abundant polymorphism.● A similar phenomenon was in the A.cylindracea YAASM1024 strains.5 fragments obtained from the amplified respectively distributed in Ⅰa,Ⅰc,Ⅰe and Ⅱb groups.The abundant polymorphism information provides a basis for the analysis of genetic differences.The result of the minimum evolution tree which based on amino acid sequence was similar as pheromone receptor nucleic acid sequence analysis.These strains could be divided into two major groups I and II,and the groups can be subdivided into seven groups from A to G Fragments located in B and D groups ware from A.cylindracea,while C,E and F groups of fragments ware from A.salicacola.A group is the largest branch,including the pheromone receptor fragments of 10 strains of A.salicacola and two fragments of A.cylindracea YAASM0594.It shows that the pheromone receptor of group A was more conservative and widely distributed between two species.Group G also contains pheromone receptors from A.cylindracea and A.salicacola.● From nucleic acid and amino acid sequence of clustering analysis of the evolutionary tree,the minimum evolution tree based on amino acid sequence was more natural,and the species distinction was more significant and sequences gathered from A.cylindracea was more concentrated.In addition,the pheromone receptor amino acid sequences from the strains of same collecting area were relatively close.Therefore,the pheromone receptor amino acid sequence analysis of the phylogenetic tree can be used to distinguish between the the A.cylindracea and A.salicacola,and as a basis for analysis of the genetic and ecological distribution.It can also provide useful information for Agrocybe genus of mating type gene structure analysis.At the same time,A.salicacola sporeless mutant strain which obtained in previous experiments as the material was studied for the reason of the basidiospore development block on,the development AFLP molecular markers associated with sporeless character which then try to convert it into SCAR marker,and the morphological and cytological compare between sporeless mutant strain and normal sporulation strain.The results show that:● For the tested strains,6 AFLP markers that consistented with the non-spore traits can be screened out through 256 primer combinations.Two of them ware successfully transformed into SCAR markers which can be carried out the non-spore traits after the screening of strains,but not widely used sporeless traits breeding work.The sequences cloned by the AFLP fragment of A.salicacola also has abundant polymorphism.● The microscopic observation results show that basidial development stopped at the sterigma stage,basidium condensed to become mononucleate.