CPC C40B 30/04 (2013.01) [C12Q 1/6806 (2013.01); C12Q 1/6869 (2013.01); C40B 40/04 (2013.01); C40B 70/00 (2013.01); C12Q 2525/205 (2013.01)] | 21 Claims |
1. A method of sequencing a peptide, comprising:
(a) incubating the peptide with a library of DNA aptamers exhibiting binding specificity toward at least one N-terminal amino acid under conditions where one or more aptamers bind specifically to at least one N-terminal amino acid of the peptide, wherein each aptamer in the library comprises a peptide binding ssDNA region and a unique barcode sequence indicative of the first binding round and the associated peptide binding ssDNA region;
(b) ligating the DNA aptamer bound to the N-terminal of the peptide onto a proximal DNA barcode construct;
(c) removing the peptide binding sequence from the DNA aptamer, thereby leaving the barcode of the DNA aptamer covalently attached to the DNA barcode construct:
(d) removing the N-terminal amino acid from the peptide to produce an N-terminal amino acid-shortened peptide;
(e) incubating the N-terminal amino acid-shortened peptide with the library of aptamers exhibiting binding specificity toward at least one N-terminal amino acid under conditions where one or more aptamers bind specifically to at least one N-terminal amino acid of the N-terminal amino acid-shortened peptide, wherein each aptamer in the library comprises a peptide binding ssDNA region and a unique barcode sequence indicative of the second binding round and the associated peptide binding ssDNA region;
(f) ligating the DNA aptamer bound to the N-terminal of the peptide onto the proximal DNA barcode construct;
(g) removing the peptide binding sequence from the DNA aptamer, thereby leaving the barcode of the DNA aptamer covalently attached to the DNA barcode construct;
(h) removing the N-terminal amino acid from the N-terminal amino acid-shortened peptide;
(i) repeating steps (a)-(d) a plurality of times to construct a chain of positional barcodes that correspond to sequential N-terminal amino acids in the peptide; and
(j) sequencing the chain of positional barcodes to thereby obtain the sequence of the peptide.
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