Protocol

Recombinant Proteins From Plants

Volume 483 of the series Methods in Molecular Biology™ pp 313-324

On-Chip Detection of Low-Molecular-Weight Recombinant Proteins in Plant Crude Extracts by SELDI-TOF MS

  • Amine M. BadriAffiliated withDépartement de phytologie, Université Laval
  • , Karine CoenenAffiliated withDépartement de phytologie, CRH/INAF, Université Laval
  • , Louis-Philippe VaillancourtAffiliated withDépartement de phytologie, CRH/INAF, Université Laval
  • , Goulet CharlesAffiliated withDépartement de phytologie, CRH/INAF, Université Laval
  • , Dominique MichaudAffiliated withDépartement de phytologie, CRH/INAF, Université Laval

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Summary

This chapter presents a general procedure for the on-chip detection and quantitation of low-molecular-weight recombinant proteins in transgenic plant crude extracts by surface-enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF MS). A protocol is first described to detect the protein of interest in crude protein extracts of transgenic plant lines, by differential protein mapping against similar extracts from a control, nontransgenic line. A complementary protocol is then presented to generate a standard curve with the SELDI system, allowing the protein to be quantified in different transgenic lines. Overall, this procedure may be carried out within a few hours, without the need for prior purification or enrichment of the recombinant protein.

Key words

Plant crude protein extracts Protein detection Protein quantitation Recombinant proteins SELDI-TOF MS Surface-enhanced laser desorption ionization time-of-flight mass spectrometry