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tRNA Probes

General information about aa - tRNAs and tRNAs

A selection of purified tRNAs is available, most of which are aminoacylated: the amber suppressor Lys-tRNAamb (recognizes the amber stop codon, UAG), yeast or E. coli Cys-tRNACys (each recognizes the UGC codon), E. coli Lys-tRNALys (recognizes the AAA codon), yeast Lys-tRNALys (recognizes the AAA codon), the E. coli initiator fMet-tRNAMetf and the yeast initiator Met-tRNAMeti (both recognize the initial AUG codon), and E. coli Phe-tRNAPhe (recognizes the UUU codon).
 
Since a wide variety of probes can be attached to the Lys or Cys side chain, we also offer these aminoacyl-tRNAs with the reactive, spectroscopic or affinity probe of your choice, while the tRNAMetf and tRNAPhe species can be modified at either the s4U-8 base or the alpha amino group of the aminoacyl-tRNA.  The proper designation for aa-tRNAs modified at the side chain amino group of Lys is Nε-(probe)-Lys-tRNA, but we abbreviate this to "εprobe-Lys-tRNA." We use "probe-S-Cys-tRNA" to designate a probe covalently attached to the Cys side chain via its sulfhydryl group.  When a probe is covalently attached to the alpha amino group of an aa-tRNA, the resulting tRNA is designated as "probe-N-aa-tRNA."

 

Yeast initiator tRNAMeti, yeast tRNALys, E. coli tRNAPhe, and E. coli tRNAMetf are chromatographically purified from unfractionated yeast or E. coli tRNA; they contain the full complement of natural modified bases.  Yeast tRNACys, E. coli tRNACys, E. coli tRNALys, and the amber suppressor tRNA (tRNAamb) are transcribed in vitro and then purified chromatographically before aminoacylation and modification; these tRNAs contain no modified bases.


After aminoacylation, all aa-tRNAs are chromatographically purified. After modification, aa-tRNAs with a fluorescent dye are chromatographically purified by HPLC, while aa-tRNAs modified with photo-reactive probes are not purified further. The extents of aminoacylation and of amino acid modification are determined directly for each aa-tRNA with a radioactive amino acid; these properties of a non-radioactive aa-tRNA are estimated by assaying a radioactive aa-tRNA prepared in parallel.