2005
DOI: 10.1016/j.femsle.2005.04.021
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Development of a sample preparation method for fungal proteomics

Abstract: Since filamentous fungi including basidiomycetous fungi possess an exceptionally robust cell wall as in microorganisms, effective extraction of intracellular proteins is a key step for fungal proteomic studies. To overcome the experimental obstacle caused by cell walls, we utilized fungal protoplasts, prepared from the brown-rot basidiomycete, Tyromyces palustris. The amount and quality of proteins extracted from the protoplast cells were much higher than that from the mycelial cells. Quantitative comparisons … Show more

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Cited by 39 publications
(22 citation statements)
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“…Several researchers (Melin et al 2002;Nandakumar et al 2003;Grinyer et al 2004;Strom et al 2005) used mechanical lysis via glass beads to liberate cytoplasmic proteins, and this approach has been reported to be more efficient than either chemical or enzymatic extraction methods (Nandakumar and Marten 2002). However, the most widely used method for extraction seems to be grinding in liquid nitrogen using mortar and pestle (Hernandez-Macedo et al 2002;Grinyer et al 2005;Shimizu and Wariishi 2005;Kniemeyer et al 2006;Yajima and Kav 2006).…”
Section: Introductionmentioning
confidence: 99%
“…Several researchers (Melin et al 2002;Nandakumar et al 2003;Grinyer et al 2004;Strom et al 2005) used mechanical lysis via glass beads to liberate cytoplasmic proteins, and this approach has been reported to be more efficient than either chemical or enzymatic extraction methods (Nandakumar and Marten 2002). However, the most widely used method for extraction seems to be grinding in liquid nitrogen using mortar and pestle (Hernandez-Macedo et al 2002;Grinyer et al 2005;Shimizu and Wariishi 2005;Kniemeyer et al 2006;Yajima and Kav 2006).…”
Section: Introductionmentioning
confidence: 99%
“…Since the fungal cell wall is exceptionally robust (Bowman and Free, 2006) and proteases are extremely resistant, cell lysis and protein extraction is challenging. Several protocols have already been published for fungal protein extraction (e.g., Nandakumar and Marten, 2002;Grinyer et al, 2004;Shimizu and Wariishi, 2005;Kniemeyer et al, 2006;Kim et al, 2008). To date, 139 intracellular proteins have been identified for various Aspergillus species (Kim et al, 2008).…”
Section: Introductionmentioning
confidence: 99%
“…As filamentous fungi possess rigid cell wall, an effective strategy has to be followed for fungal proteomics study. Shimizu and Wariishi has devised an appropriate method for lysing the fungal cell wall by generating protoplasts where 2DE patterns were better visualised than proteins isolated from disrupting the fungal cell wall using SDS extraction [35]. The protein extracted is subjected to Tri Chloro Acetic Acid (TCA) precipitation for purification to minimise problems due to contaminants during isoelectric focusing [36].…”
Section: Protein Isolationmentioning
confidence: 99%