Primer set details

ORGANISM
Complete name : Phytophthora spp.
Database ID : 205
INTERNET RESOURCES
Google Wikispecies, the free species directory Wikipedia, The Free Encyclopeda
NCBI Taxonomy Browser CABI Index Fungorum CBS Anamorph/Teleomorph

PRIMER SET (5’→3′)
Forward Primer Select to Blast
Name : Ph2 Sequence :
Reverse Primer
Name : ITS4 Sequence :
TARGET DNA
ITS region
PCR TECHNIQUE
Nested PCR
REMARKS
First-Round primers: ITS5-ITS4 (White et al., 1990)
REFERENCE
Authors : Ippolito A., Schena L., Nigro F.
Year : 2002
Title : Detection of Phytophthora nicotianae and P. citrophthora in citrus roots and soils by nested PCR
Journal : European Journal of Plant Pathology
Volume : 108
Pages : 855–868
Abstract : The polymerase chain reaction (PCR) was used for the specific detection of Phytophthora nicotianae and P. citrophthora in citrus roots and soils. Primers were based on the nucleotide sequences of the internal transcribed space regions (ITS1 and ITS2) of 16 different species of Phytophthora. Two primer pairs, Pn5B-Pn6 and Pc2B-Pc7, were designed specifically to amplify DNA from P. nicotianae and P. citrophthora, respectively. Another primer pair (Ph2-ITS4) was designed to amplify DNA from many Phytophthora species. All primer pairs were assessed for specificity and absence of cross-reactivity, using DNA from 118 isolates of Phytophthora and 82 of other common soil fungi. In conventional PCR, with a 10-fold dilution series of template DNA, the limit of detection was of 1 pg/microliter DNA for all the primer pairs (Ph2-ITS4, Pn5B-Pn6, and Pc2B-Pc7). In nested PCR, with primers Ph2-ITS4 in the first round, the detection limit was of 1 fg/microliter for both the primer sets (Pn5B-Pn6 and Pc2B-Pc7). Simple, inexpensive and rapid procedures for direct extraction of DNA from soil and roots were developed. The method yielded DNA of a purity and quality suitable for PCR within 2-3 h. DNA extracted from soil and roots was amplified by nested PCR utilizing primers Ph2-ITS4 in the first round. In the second round the primer pairs Pn5B-Pn6 and Pc2B-Pc7 were utilized to detect P. nicotianae and P. citrophthora, respectively. Comparison between the molecular method and pathogen isolation by means of a selective medium did not show any significant differences in sensitivity.
SUBMITTER
Dr. Stefano Ghignone (Sistem Administrator)
stefano.ghignone@ipsp.cnr.it
Consiglio Nazionale delle Ricerche (CNR)
Istituto per la Protezione delle Piante, Sez. Torino
V.le P.A. Mattioli 25, I-10125 Turin
Italy
Record creation : 2004-04-13 Last record update : 2004-04-13

* Disclaimer *

No guarantee can be given for the correctness of the data presented.
Although the DNA sequences were reviewed for accuracy,
original pubblications should be consulted to verify their composition.

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