Fusion protein aptamer screening method and kit
A nucleic acid aptamer and fusion protein technology, applied in the field of genetic engineering, can solve the problems of high screening cost, complicated operation, single target, etc., and achieve the effect of improving high efficiency, small adsorption effect, and good biocompatibility
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Embodiment 1
[0050] Example 1. The method for screening the expression of Strep-TagII in Escherichia coli cells using the fusion protein nucleic acid aptamer screening kit
[0051] (1) Centrifuge 100 µL of Escherichia coli fusion protein expressing bacteria solution and 100 µL of non-expressing protein bacterial solution at 4°C, 12,000 r / min for 30 minutes, discard the supernatant, and incubate with 100 µL agar magnetic beads at 37±0.3°C for 30 minutes, wash Wash once with buffer solution, add protein blocking solution and seal at 37±0.3°C for 1 hour, remove the supernatant, wash once with washing buffer solution, and magnetically separate to obtain positive sieve tubes (magnetic bead E. coli fusion expression protein bacterial solution) And anti-sieve tube (magnetic beads E. coli non-expressing protein bacteria liquid).
[0052] (2) Forward binding: In the first round of screening, the random oligonucleotide library (ssDNA library) was dissolved in binding buffer, heated at 95°C for 5 min...
Embodiment 2
[0058] Example 2, the method of screening the expression of GST by Pichia pastoris using the fusion protein nucleic acid aptamer screening kit
[0059] (1) Centrifuge 100 µL Pichia fusion protein expressing bacteria solution and 100 µL non-expressing protein bacterial solution at 4°C, 12,000 r / min for 30 minutes, discard the supernatant, and incubate with 100 µL agar beads at 37±0.3°C for 30 minutes, Wash once with washing buffer, add protein blocking solution and seal at 37±0.3°C for 1 hour, remove the supernatant, wash once with washing buffer, and magnetically separate to obtain positive screen tubes (Pichia magnetic beads fusion expressing proteobacteria) liquid) and anti-sieve tube (Pichia magnetic beads non-expressing protein bacteria liquid).
[0060] (2) Forward binding: In the first round of screening, the random oligonucleotide library (ssDNA library) was dissolved in binding buffer, heated at 95°C for 5 minutes and cooled rapidly in ice water for 10 minutes, then ad...
Embodiment 3
[0066] Example 3, the method for screening the expression of His-tag by baculovirus using fusion protein nucleic acid aptamer screening kit
[0067] (1) Centrifuge 100 µL of baculovirus fusion protein-expressing bacterial solution and 100 µL of non-expressing protein bacterial solution at 4°C, 12,000 r / min for 30 minutes, discard the supernatant, and incubate with 100 µL of agar beads at 37±0.3°C for 30 minutes. Wash once with washing buffer, add protein blocking solution and seal at 37±0.3°C for 1 hour, remove the supernatant, wash once with washing buffer, and magnetically separate to obtain positive screen tubes (magnetic bead baculovirus fusion expressing proteobacteria) solution) and anti-sieve tube (magnetic bead baculovirus non-expressing protein bacteria solution).
[0068] (2) Forward binding: In the first round of screening, the random oligonucleotide library (ssDNA library) was dissolved in binding buffer, heated at 95°C for 5 minutes and cooled rapidly in ice water...
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