Molecular clustering of Pycnoporus strains

from various geographic origins and isolation of monokaryotic strains for laccase hyperproduction.

Original article by: Anne Lomascolo, Jean-Luc Cayol , Marjolaine Roche , Lin Guo , Jean-Luc Robert,Eric Record , Laurence Lesage - Meessen , Bernard Ollivier, Jean-Claude Sigoillot and Marcel Asther

By: Adrián García

INTRODUCTION

• Several species among the genus Pycnoporus have shown the capacity to produce the enzyme laccase, an enzyme widely used for industrial purposes. (biobleaching, wastewater treatment, etc.)
• The species P. Sanguineus has been studied and is considered a very efficient producer of the enzyme. 
•  A common phenomenon on the fungal life cycle, is the formation of dikaryotic mycelium, which contains 2 different nuclei, instead of a monokaryotic mycelium, which contains only 1 nucleus.

Life cycle

PROCESS

• Research is made to isolate the best laccase producing strain from wild backgrounds and/or from pre-existing collections.

• In order to fulfil this task, the strains were screened using different techniques to find the superior strain.
  • These techniques include genetic sequencing, and monokaryotic strain isolation.

MATERIALS AND METHODS

• Strains:
  • 8 wild, tropical P. Sanguineus strains collected from Chinese provinces.
  • 20 P. cinnabarinus,  P. coccineus and P. sanguineus strains from international pre-existing collections.
  • Strain G05 (tropical Chinese) was tested with different inducers and with a different culture medium containing Tween 80.

Strain culture

Laccase production

• Maltose medium with ferulic acid as inducer 

* Other inducers were tested

Monokaryon isolation of dikaryotric strains

DNA

isolation

PCR

amplification

DNA

sequencing 

 ITS1-5.8S-ITS2

Laccase activity assay

• ABTS

Monokaryotic strain culture

RESULTS

• Laccase production:

  • P. sanguineus proved to be a better enzyme producer than P. cinnabarinus and  P. coccineus. 

  • P. sanguineus G05.10 produced up to 71000 U/L

  • Best culture medium contained:

    • Tween 80

    • 2,5-xylidine

    • Maize bran

• Monokaryotic isolation and laccase production:
  • Monokaryotic isolation from dikaryotic parents increased enzyme production in most cases.

RESULTS

• Strain origin and phylogenetic relations:​

  • Chinese Pycnoporus strains were better laccase producers than the other analysed strains. Using monokaryotic strains through isolation, improves laccase production. 

  • Geographical origin proved to be a stronger determinant of phylogenetic relations, over taxonomical position.

APPLICATION & POTENTIAL

• Strain origin and phylogenetic relations:

  • Geographical origin proved to be a stronger determinant of phylogenetic relations, over taxonomical position.

  • Chinese Pycnoporus strains were better laccase producers than the other analysed strains. Using monokaryotic strains through isolation, improves laccase production. 

THANKS!

All data was retrieved from the article 

Molecular clustering of Pycnoporus strains from various

geographic origins and isolation of monokaryotic strains for

laccase hyperproduction, by: 

 Anne Lomascolo, Jean-Luc Cayol , Marjolaine Roche , Lin Guo , Jean-Luc Robert,Eric Record , Laurence Lesage - Meessen , Bernard Ollivier, Jean-Claude Sigoillot and Marcel Asther