Myrtle Rust Fungus

Myrtle rust is an exotic rust fungus originating from South American region, detected for the first time in Australia (New south wales) on 22nd April 2010, the fungus was found growing on syncarpia glomulifera, callistemon viminalis and agonis fluxuosa plants. The infected plants can be easily identified from powdery bright orange-yellow or yellow spores on fruits, buds, leaves and shoots.

As the rust fungus is considered to be a biosecurity threat, a state emergency response program was initiated by the Australian government, which conducted surveys in NSW in 2010, the data collected from the surveys shows that the fungus has already been recorded on 107 host species in 30 genera.

Understanding the genome:

Advanced molecular biology techniques

The key enzyme in the process is DNA polymerase that forms the PCR product by linking individual nucleotide bases (adenine, guanine, cytosine and thymine). Short DNA fragments with sequence complimentary to target DNA are called primers; these specify the DNA sequence that has to be amplified. The PCR reaction mixture is made in a 96 well plate or test tube and placed in a thermal cycler.

The thermal cycler is a machine used to amplify DNA by altering the temperature in repeated cycles using three precise pre programmed steps:

Denaturation – By increasing the temperature in the thermal cycler the DNA is denatured creating separate single stranded DNA

The DNA copies are doubled with each repetition of the three steps. The PCR products are usually analysed using Agarose gel electrophoresis methods, which separates the DNA fragments based on size.

A study carried out at the University of Sydney under the National Myrtle Rust Transition to Management (T2M) Program used polymorphic satellite markers technique to understand the genetic variability of Myrtle rust isolates. The amplification of the satellite markers was done using PCR with specific controls.

Reactions were performed in a 96-well DNA thermo cycler (Eppendorf Mastercycler, Germany) using the following reaction mixture:

• 2.0 μl of genomic DNA (10 ng/μl)

• 1.5 μl of 10x PCR buffer (NH4 Reaction buffer, Bioline)

• 1.5 μl of dNTPs (0.2 mM)

• 0.9 μl of 50 mM MgCl2 (Bioline)

• 0.9 μl of each forward and reverse primer (2 mM)

• 0.15 μl (5 u/μl) of Taq DNA (Bioline, Australia)

• 7.15 μl of