Comparative Analysis And Population Structure Of Sexual Offsprings In Phytophthora Infestans

Potato late blight?PLB?caused by Phytophthora infestans?P.infestans?is one of the most devastating diseases in potato around the world.In this study,we collected P.infestans from potato main planting region in Kunming,Yunnan Province,then isolated and purified it.Besides,after determining its species using morphology and ITS sequence,the chromosome ploidy,mating type,mitochondrial haplotype,virulence and metalaxyl sensitivity were determined.Selecting 5 isolates,sexual reproduction F₁ progenies were obtained by using the improved oospore germination method.And then the phenotype?mating type,virulence and metalaxyl sensitivity?and genotype?chromosome ploidy,mitochondrial haplotype,SSR marker?of F₁ progenies were detected,analyzed the relationship between phenotype and genotype.Based on results above,sexual reproduction F₁ progenies that have 4 mating types?A1,A2,A1A2,SF?induced by a pair P.infestans parents were used for the whole genome resequencing and comparative analysis.The main results were:1.In the middle of July 2017,19 P.infestans collected from different species potato in Xundian and Songming in Kunming,Yunnan Province,were isolated,purified and determined.By paired culture method,3 of them were A1 mating type,12 of them were A2 mating type,1 of them was A1A2 mating type,3 of them were SF.By detecting mitochondrial DNA of those 19 P.infestans,all of them were?a,which was dominant population in Yunnan Province.2.Using low temperature to induce oospore germinationby 6 pair of parents,the optimal of freezing treatment for 24 h is the best condition,the oospore germination rate may reach to 5.80%±0.2%.In addition,all the F₁ progenies were diploid and had the same mitochondrial haplotype with parents which implied that it had not recombined.The result also found that there was a segregation in F₁ progenies toward mating type,metalaxyl sensitivity and virulence.The mating type separation ratio is 16:5:17:22?A1:A2:A1A2:SF?,the sensitivity ratio to metalaxyl is 1:14?R:S?,and the virulence separation ratio is 5:19?R:S?,which was obviously abide by mendelian separation rules.Genotype analysis by 8 pair SSR markers showed that at the genetic similarity coefficient down to 0.95,all isolates could be divided into 14 genotypes and 6 clades,in which S1was dominant population,accounting for 61.67%.The relationship analysis revealed significantly correlation between genotypes and phenotypes?R2=0.6667?.3.4 segregant populations based on mating type?BSA1,BSA2,BSA3 and BSA4?,were selected for Illumina resequencing PE150?10X?,assembly,annotation and BSA analysis by CLC genomic workbench 11.0.1,with a reference genome of T30-4?A1mating type?obtained in NCBI.It was found that there was a total SNP of 528712 in the BSA4 segregant populations.Furthermore,BSA4 has the most Indels of 20849,while BSA2 has the least Indels of 17746.BSA4 has the most SV of 6613,while BSA2 has the least SV of 5097.The comparison each other populations found that the number of SNPs between BSA1 and BSA2 was the highest,reached 38551,the least being BSA2 and BSA4,only 19630.From the above results,it was concluded that isolates which were collected in Xundian and Songming,Kunming City,Yunnan Province in 2017,acquisition 19 strains of P.infestans,P.infestans in four?A1,A2,A1A2 and SF?mating types,but A1 mating type of is less than other,and all of 19 strains sharing?a mitochondrial haplotype;the germination of oospores was improvement by low temperature treatment.Both,phenotype and genotype were also analyzed among sexual offsprings;In addition,the genome was resequenced,assembled and annotated from 4 segregation populations,to further understanding the functional genes or mating type linked locus by comparative genomics analysis.