Abstract
The generation of peptide from messenger RNA (mRNA) provides a convenient source for current proteomic analysis. Intron-free mRNA possessing adenine-uracil-guanine (AUG) start codons can be translated into labeled or unlabeled peptides under a predetermined reticulocyte lysate condition. In conjunction with RNA-polymerase cycling reaction (see Fig. 1; RNA-PCR), full-length gene transcripts can be unlimitedly amplified for protein/peptide synthesis in vitro (1). Many commercialized in vitro translation systems provide a cap nucleotide, which can be added to the 5′ end of the amplified poly(A+) RNAs during the transcription step of RNA-PCR. Totally resembling mRNAs, the capped poly(A+) RNAs can be used to synthesize proteins/peptides with labeling and may help the functional analysis of protein activity if they fold correctly.
An illustration of using RNA-PCR procedure for peptide library generation. The mRNA generated in step d can be capped by a methylated nucleotide such as P1-5′-(7-methyl)-guanosine-P3-5′-adenosine/guanosine-triphosphate for in vitro translation.
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© 2003 Humana Press Inc., Totowa, NJ
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Lin, SL. (2003). Peptide Library Construction from RNA-PCR-Derived RNAs. In: Ying, SY. (eds) Generation of cDNA Libraries. Methods in Molecular Biology™, vol 221. Humana Press. https://doi.org/10.1385/1-59259-359-3:289
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DOI: https://doi.org/10.1385/1-59259-359-3:289
Publisher Name: Humana Press
Print ISBN: 978-1-58829-066-3
Online ISBN: 978-1-59259-359-0
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