Oligo DNA barcodes were designed based on the LIBRA-seq protocol (9 (link)). In brief, a 15 nt DNA barcode was inserted into the following sequence: 5’-CCTTGGCACCCGAGAATTCCA-Barcode-CCCATATAAGA*A*A-3’. The barcodes used in this study were: TCCTTTCCTGATAGG(Spike), GCTCCTTTACACGTA(RBD). All the oligo barcodes were synthesized with 5’ amino modifier C6 and purified by HPLC (Integrated DNA Technologies; IDT). After purification, AviTag labeled proteins were biotinylated using the BirA biotin-protein ligase reaction kit (Avidity LLC, cat no. BirA500). The barcode oligos were linked to biotinylated S-ecto or RBD using the Solulink Protein-Oligonucleotide Conjugation Kit (TriLink cat no. S-9011) according to manufacturer’s instructions.
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Oligo DNA Barcoding for Protein Analysis
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Biotin ligase
Hplc
Oligonucleotide
Oligos
Protein
Corresponding Organization :
Other organizations : The University of Texas Southwestern Medical Center, The University of Texas Medical Branch at Galveston, Howard Hughes Medical Institute
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Variable analysis
independent variables
- DNA barcode sequence design
- Insertion of 15 nt DNA barcode into the sequence
- Barcodes used: TCCTTTCCTGATAGG (Spike), GCTCCTTTACACGTA (RBD)
- Modification of oligo barcodes with 5' amino modifier C6
- Purification of oligo barcodes by HPLC
- Biotinylation of AviTag labeled proteins using BirA biotin-protein ligase reaction kit
- Linking of barcode oligos to biotinylated S-ecto or RBD using Solulink Protein-Oligonucleotide Conjugation Kit
- Not explicitly mentioned
- Not explicitly mentioned
- Positive control: Not mentioned
- Negative control: Not mentioned
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