title: Mitochondrial DNA polymorphisms in Phytophthora infestans: new haplotypes are identified and re-defined by PCR.
abstract: Polymorphisms of mitochondrial DNA (mt-DNA) are particularly useful for monitoring specific pathogen populations like Phytophthora infestans. Basically type I and II of P. infestans mt-DNA were categorized by means of polymorphism lengths caused by an ~2 kb insertion, which can be detected via restriction enzyme digestion. In addition genome sequencing of haplotype Ib has been used as a simple Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) method to indirectly identify type I and II alterations through EcoR I restriction enzyme DNA fragment patterns of the genomic P4 area. However, with the common method, wrong mt-DNA typing occurs due to an EcoR I recognition site mutation in the P4 genomic area. Genome sequencing of the four haplotypes (Ia, Ib, IIa, and IIb) allowed us to thoroughly examine mt-DNA polymorphisms and we indentified two hypervariable regions (HVRs) named HVRi and HVRii. The HVRi length polymorphism caused by a 2 kb insertion/deletion was utilized to identify mt-DNA types I and II, while another length polymorphism in the HVRii region is caused by a variable number of tandem repeats (n = 1, 2, or 3) of a 36 bp sized DNA stretch and was further used to determine mt-DNA sub-types, which were described as R(n = 1, 2, or 3). Finally, the P. infestans mt-DNA haplotypes were re-defined as IR(1) or IIR(2) according to PCR derived HVRi and HVRii length polymorphisms. Twenty-three isolates were chosen to verify the feasibility of our new approach for identifying mt-DNA haplotypes and a total of five haplotypes (IR(1), IR(2), IR(3), IIR(2) and IIR(3)) were identified. Additionally, we found that six isolates determined as type I by our method were mistakenly identified as type II by the PCR-RFLP technique. In conclusion, we propose a simple and rapid PCR method for identification of mt-DNA haplotypes based on sequence analyses of the mitochondrial P. infestans genome.
authors: College of Plant Protection, Agricultural University of Hebei, Baoding 071001, China.
authors: Zhi-Hui
authors: ZH
authors: Yang
authors: Ming-Xing
authors: MX
authors: Qi
authors: Yu-Xuan
authors: YX
authors: Qin
authors: Jie-Hua
authors: JH
authors: Zhu
authors: Xiu-Mei
authors: XM
authors: Gui
authors: Bu
authors: B
authors: Tao
authors: Xiao-Hu
authors: XH
authors: Xu
authors: Fu-Guang
authors: FG
authors: Zhang
mesh_terms: D017931:DNA Primers / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics*; D006239:Haplotypes; D055750:Phytophthora infestans / Q000235:genetics* / Q000302:isolation & purification; D010935:Plant Diseases / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D012150:Polymorphism, Restriction Fragment Length; D017422:Sequence Analysis, DNA; D011198:Solanum tuberosum / Q000382:microbiology
chemical_list: D017931:DNA Primers; D004272:DNA, Mitochondrial
keywords: Mitochondrial haplotypes; Oomycetes; Phytophthora infestans; Potato late blight