Analysis of molecular variability among isolates of Aspergillus flavus by PCR-RFLP of the ITS regions of rDNA

• Autorzy: Mohankumar M. , Vijayasamundeeswari A. , Karthikeyan M. , Mathiyazhagan S. , Paranidharan V. , Velazhahan R.

Warianty tytułu

PL

Analiza molekularnej zmienności izolatów Aspergillus flavus przy wykorzystaniu regionów rDNA i techniki PCR-RFLP

• Języki publikacji: EN

Abstrakty

EN

A total of seventeen isolates of Aspergillus flavus from maize were collected from different agro-ecological zones of Tamil Nadu, India. The isolates were tested for their ability to produce aflatoxin B1 (AFB1) in vitro by indirect competitive enzyme-linked immunosorbent assay (ELISA). The amount of AFB1 produced by the isolates of A. flavus ranged from 1.9 to 206.6 ng/ml. Among the various isolates of A. flavus, the isolate AFM46 produced the highest amount of AFB1. DNA was extracted from A. flavus isolates and their molecular variability was investigated by using restriction fragment length polymorphism (RFLP) analysis of the PCR-amplified internal transcribed spacer (ITS) regions of ribosomal DNA. PCR amplification with ITS1 and ITS4 primers resulted in the amplification of a product of approximately 600 bp. Digestion of the PCR products with the restriction enzymes EcoRI, HaeIII and TaqI produced fragments of different sizes. Analysis of the genetic coefficient matrix derived from the scores of RFLP profiles showed that minimum and maximum per cent similarities among the tested A. flavus strains ranged from 0 to 88%. Cluster analysis using the unweighted pair-group method with arithmetic average (UPGMA) clearly separated the isolates into five groups (group I–V) confirming the genetic diversity among the A. flavus isolates from maize.

PL

Zebrano ogółem siedemnaście izolatów Aspergillus flavus z kukurydzy, z różnych stref agro-ekologicznych Tamil Nadu, Indie. Badano zdolność izolatów do wytwarzania aflatoksyny B1 (AFB1) in vitro, wykorzystując pośredni test ELISA (enzymatyczny test immunosorbcyjny). Ilość wytworzonego AFB1 przez izolaty A. flavus wahała się w granicach od 1,9 do 26,6 mg/ml. Spośród różnych izolatów A. flavus, izolat AFM146 wytwarzał największą ilość AFB1. Ekstrahowano DNA z izolatów A. flavus i określano ich molekularną zmienność, wykorzystując analizę poliformizmu długości fragmentów restrykcyjnych (RLFP) regionów rybosomalnego DNA. Amplifikowanie DNA przy wykorzystaniu starterów ITS1 i ITS4, spowodowało amplifikację produktu wynoszącą, w przybliżeniu, 600 bp. Strawienie produktów PCR enzymami restrykcyjnymi EcoRI, HaeIII i TaqI, dało, w wyniku, fragmenty o różnych wartościach genetycznego współczynnika matrix, pochodzącego z ocen. Analiza profilów DNA wykazała, że minimalny i maksymalny procent podobieństwa wśród testowanych ras A. flavus wynosił od 0 do 88. Analiza skupień metodą para-grupa przy wykorzystaniu średniej arytmetycznej (UPGMA) wyraźnie dzieliła izolaty na pięć grup (grupy I–IV), potwierdzając genetyczną różnorodność wśród izolatów A. flavus z kukurydzy.

Słowa kluczowe

EN

molecular variability; isolate; Aspergillus flavus; rDNA region; PCR-RFLP method; aflatoxin B1; genetic variability; internal trancribed spacer polymerase chain reaction; RFLP analysis

• Wydawca: -
• Czasopismo: Journal of Plant Protection Research
• Rocznik: 2010
• Tom: 50
• Numer: 4
• Opis fizyczny: p.446-451,fig.,ref.

Twórcy

autor

Mohankumar M.

• Department of Plant Pathology, Centre for Plant Protection Studies, Tamil Nadu Agricultural University, Coimbatore- 641 003, Tamil Nadu, India

autor

Vijayasamundeeswari A.

autor

Karthikeyan M.

autor

Mathiyazhagan S.

autor

Paranidharan V.

autor

Velazhahan R.

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