TAM-ChIP Multiplex Protein Isolation
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Solution Overview
Problem
Current chromatin immunoprecipitation (ChIP) techniques are limited by high sample size requirements, inability to analyze multiple proteins concurrently, and the need for ultra-high affinity antibodies, which restricts their application in fields like stem cell research and high-throughput screening.
Innovation Solution
The development of Transposase-Assisted Multi-analyte Chromatin ImmunoPrecipitation (TAM-ChIP) method, which uses antibody-transposome complexes to tag and isolate chromatin-associated nucleic acids, allowing for simultaneous analysis of multiple proteins and reducing sample requirements through transposition-based library preparation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If traditional chromatin immunoprecipitation (ChIP) techniques are used, then protein-chromatin interactions can be analyzed, but large sample sizes are required and multiple proteins cannot be analyzed concurrently
Solution Approach 1:
The invention segments the analysis process by using unique bar code sequences associated with each antibody-transposome complex. This allows multiple proteins to be analyzed concurrently in a single reaction mixture, with each protein's associated DNA fragments tagged with distinct bar codes that enable later identification and separation of the different protein targets from the same chromatin sample.
Solution Approach 2:
The invention creates a universal platform where antibody-transposome complexes can simultaneously target multiple different proteins in the same chromatin sample. The transposome component provides universal functionality for DNA tagging and bar code incorporation, while the antibody component provides specific targeting capability, enabling one system to perform multiple protein analysis functions concurrently.
2Adaptability or versatility
If traditional ChIP techniques are used, then protein-chromatin interactions can be studied, but ultra-high affinity antibodies are required
Solution Approach 1:
The invention introduces transposome complexes as intermediaries between the antibody and the DNA. Instead of requiring the antibody to directly pull down chromatin with ultra-high affinity, the antibody binds to the protein of interest, and the transposome component mediates the tagging of associated DNA fragments with bar code sequences. This intermediary mechanism reduces the stringency requirements for antibody-chromatin binding.
Solution Approach 2:
The invention replaces the mechanical immunoprecipitation process with a biochemical transposition-based tagging system. Instead of relying on physical pulling down of chromatin by antibodies (which requires ultra-high affinity), the system uses enzymatic transposition to insert bar-coded oligonucleotides at the sites where antibodies are bound, substituting a biochemical mechanism for a mechanical one and thereby reducing antibody affinity requirements.
3Productivity
If traditional ChIP techniques are used, then DNA isolation can be performed, but labor-intensive library preparation is required
Solution Approach 1:
The invention performs preliminary action by incorporating bar code sequences and adapter sequences into the DNA fragments during the transposition step itself, rather than requiring separate library preparation steps later. The transposome complexes pre-load the necessary sequencing adapters and bar codes onto the DNA fragments as they are being tagged, so that when the DNA is extracted and amplified, it is already in a format ready for high-throughput sequencing, eliminating time-consuming library preparation steps.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
TAM-ChIP enables rapid, streamlined analysis of protein-chromatin interactions with reduced sample size needs, eliminating the need for immunoprecipitation and labor-intensive library preparation, thereby expanding the range of analyzable proteins and enhancing the efficiency of epigenetic research.
Implementation Method 1
The development of Transposase-Assisted Multi-analyte Chromatin ImmunoPrecipitation (TAM-ChIP) method, which uses antibody-transposome complexes to tag and isolate chromatin-associated nucleic acids
Implementation Method 2
adding at least one antibody-oligonucleotide conjugate comprising an extraction moiety, allowing said antibody(ies) to locate at its/their target protein(s) in said chromatin fragments
Implementation Method 3
extracting the nucleic acid so tagged using the extraction moiety
Data Source
AI summary
The invention provides novel methods and materials for genetic and genomic analysis using single or multiplex isolation of protein-associated nucleic acids, including transposase-assisted chromatin immunoprecipitation (TAM-ChIP) and antibody-oligonucleotide proximity ligation. These methods comprise tagging and isolating chromatin or other protein-associated nucleic acids and using antibody-oligonucleotide complexes that recognize the proteins associated with such nucleic acids.