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Detection of the Recombinant Proteins in Single Transgenic Microbial Cell Using Laser Tweezers and Raman Spectroscopy
journal contribution
posted on 2007-12-15, 00:00 authored by Changan Xie, Nhu Nguyen, Yong Zhu, Yong-qing LiLaser tweezers Raman spectroscopy (LTRS) has been
used for the rapid detection of recombinant somatolactin
protein produced in single Escherichia coli bacteria and
Pichia pastoris yeast cell in the current study. A cDNA
sequence encoding mature peptide of zebrafish somatolactin β was inserted into two different expression vectors
and transfected into E. coli or P. pastoris yeast cells. We
measured Raman spectra of single E. coli cells at different
culture times following the induction with isopropyl β-d-1-thiogalactopyranoside, from which the amount of the
generated somatolactin proteins was obtained by the
projection of the entire cell's spectrum onto the spectrum
of the pure somatolactin proteins or the dot product
between these two spectral vectors. We found that the
intensity of the somatolactin β protein-associated spectra
from single E. coli cells increased as the function of the
culture time, which correlates with the accumulation of
recombinant proteins inside the cells. This spectral
observation was supported by evidence obtained by
conventional methods of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting analyses.
The increased intensities of recombinant protein-associated Raman bands were also observed in another expression system, P. pastoris yeast cells. These findings
demonstrate that the LTRS is a useful method for rapid
sensing of recombination production in single host microorganism in vivo.