Allelic variation at a hypervariable compound microsatellite locus in the ascomycete Ascochyta rabiei

被引:25
作者
Geistlinger, J
Weising, K
Kaiser, WJ
Kahl, G
机构
[1] UNIV FRANKFURT, DEPT BIOL, D-60439 FRANKFURT, GERMANY
[2] WASHINGTON STATE UNIV, AGR RES SERV, USDA, PULLMAN, WA 99164 USA
来源
MOLECULAR AND GENERAL GENETICS | 1997年 / 256卷 / 03期
关键词
locus-specific microsatellite analysis; microsatellite-primed PCR; RFLP analysis; mutagenesis; phytopathogenic fungi;
D O I
10.1007/s004380050573
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The genome of the fungal chickpea pathogen Ascochyta rabiei was screened for polymorphisms by microsatellite-primed PCR. While ethidium-bromide staining of electrophoretically separated amplification products showed only limited polymorphism among 24 Tunisian A, rabiei isolates, Southern hybridization of purified PCR fragments to restriction digests of fungal DNA revealed polymorphic DNA fingerprints. One particular probe that gave rise to a hypervariable single-locus hybridization signal was cloned from the Syrian isolate AA6 and sequenced. It contained a large compound microsatellite harbouring the penta-and decameric repeat units (CATTT)(n), (CATTA)(n), (CATATCATTT)(n) and (TATTT)(n),. We call this locus ArMS1 (Ascochyta rabiei microsatellite 1). Unique flanking sequences were used to design primer pairs for locus-specific microsatellite amplification and direct sequencing of additional ArMS1 alleles from Tunisian and Pakistani isolates. A high level of sequence variation was observed, suggesting that multiple mutational mechanisms have contributed to polymorphism. Hybridization and PCR analyses were performed on the parents and 62 monoascosporic F-1 progeny derived from a cross between two different mating types of the fungus. Progeny alleles could be traced back to the parents, with one notable exception, where a longer than expected fragment was observed. Direct sequencing of this new length allele revealed an alteration in the copy number of the TATTT repeat [(TATTT)(53) to (TATTT)(65)], while the remainder of the sequence was unchanged.
引用
收藏
页码:298 / 305
页数:8
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