Cloning And Analysis Of Mating Type Gene Fragments In Setosphaeria Turcica

Northern corn leaf blight(NCLB)is a severe disease on cron.The pathogen has two life styles,that is asexual life style and sexual life style.In the sexual life style the pathogen is called Setosphaeria turcica,belonging to heterothallic ascomycetes.In the sexual life style,the isolates can be individed into two mating types:the "A" mating type and the "a" mating type.Isolates of opposite mating types can be induced to generate ascus when growing on special medium.In filamentous fungus the mating process was controlled by a pair of allele genes called mating type gene,named as MAT1-1 and MAT1-2,which were contained in opposite isolates respectively.The translated MAT1-1 contains a "α" DNA binding domain while the translated MAT1-2 contains a HMG-BOX domain,these two genes belong to different DNA binding protein families.This research used the isolates 40(mating type A),01-23(mating type a)and 03-08 (mating type Aa)as materials.The genome DNA of these isolates were extracted using CTAB method and the fragments of mating type genes were cloned using degenerated primers.A fragment of 859 base pair in length was cloned from isolate 01-23,containing a "α" domain.A fragment of 238 base pair in length was cloned from isolate 40,containing a HMG-BOX domain.We optimized the condition of PCR reacetion and established a practical method in mating type identification of Setosphaeria turcica.This method was faster and more accurate than the traditional confronting method.Ascus was generated in the confronting experiment of 03-08 against 03-08, suggesting the isolate of self-fertilization.Both fragments of mating type gene were cloned from isolate 03-08 by PCR amplification,suggesting that the self fertilization of isolate 03-08 having its genetic basis.