Genetic Diversity And RDNA ITS Sequence Analysis Of The Dominant Pathogens (Alternaria Spp.) Of Wheat Black Point In Henan

The genetic diversity of 43 Alternaria spp.isolates from the wheat black point of six representative areas in Henan Province was analyzed by RAPD.151 RAPD markers were obtained,which were amplified by 17 random primers,with 100 %polymorphisms.It showed that rich genetic diversity of the pathogens of wheat black point was existed in Henan Province.The results of NTSYS cluster analysis showed that Alternaria alternata and A.tenuissima were the main pathogens of wheat black point,and the similarity coefficients were ranged from 0.62 to 0.92.Most of the isolates from the same area were clustered together and showed close genetic relationships.Some isolates from different areas were also clustered together with similarity.The results of genetic diversity analysis to the isolates had basically proved the morphological identification.8 samples of Alternaria alternata were selected from the isolates,and their genomic DNAs were extracted.The ribosomal DNA internal transcribed spacer(ITS) region of the genomic DNAs was amplified and sequenced.Nucleotide sequence analysis showed that this 8 isolates have high homology with the standard strain 34-016 and other 32 isolates which come from different places and different sources in GenBank. All the isolates did not have the regionally characters,however,some isolates which come from one place have more same base mutation than that from different places; One isolate YJ949-1 of the 9 A.alternata isolates had one same base mutation with that from Italy and the homology is 98%.So rDNA-ITS ITS1-5.8S-ITS2 sequences have certain mutation based on the relative conservation,which is the important evidence in the classification,identification and phylogenetic analyses of some epiphyte.According to the sequence analyse of A.alternata isolates from many areas of the world,the results showed that rDNA ITS 1-5.8S-ITS2 can not be the criterion to classify the different isolates from different geographical zones and sources. In order to study phylogenetic relationship of the pathogens of wheat black point in Henan,9 samples of A.tenuissima which is also the pathogen of wheat black point were analyzed.The ribosomal DNA internal transcribed spacer(ITS) region of the genomic DNAs was amplified and sequenced,the homology of the 17 isolates was 99%-100%.Compared with the 9 species of Alternaria's sequence from GenBank, the resμLt shows that all the isolates have high homology and the homology of the 6 small-spored species A.alternata,A.tenuissima,A.mali,A.gaisen,A.citri and A. arborescens is 98%-100%.But the three large-spored species A.radicina,A.porri and A.solani are separated clearly from the 6 small-spored species.According to the sequence analysis of 58 isolates in Alternaria spp.,the isolates of the same or similar species have high conservation no matter the host or region.Compared with the small-spored species,3 large-spored species have special base mutation in the site of 97-171,365-402,443-490.So these special sequences can be important evidence in the classification,identification,molecular marker and phylogenetic analyse of some certain species.The ITS1-5.8S-ITS2 sequence analysis can only prove the morphological resμLt of the pathogens of wheat black point,but it can not be used in the classification of the similar species of Alternaria spp..