ChIP-seq of BpTCP20-overexpressing plants
Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) was performed to identify genome-wide binding sites of BpTCP20. Overexpressing BpTCP20 plants were treated with formaldehyde cross-linking to fix protein-DNA interactions. Subsequently, nuclei were isolated and chromatin was fragmented into 200–500 bp by sonication. Flag antibody was used for specific immunoprecipitation of BpTCP20-DNA complexes, with Input and negative controls set to ensure result reliability. After reverse cross-linking and purification, the precipitated DNA was used for ChIP-seq library construction and high-throughput sequencing. Finally, bioinformatics analyses (including sequence alignment, Peak calling, annotation, and Motif analysis) were performed to identify the precise binding sites of BpTCP20 genome-wide.
Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) was performed to identify genome-wide binding sites of BpTCP20. Overexpressing BpTCP20 plants were treated with formaldehyde cross-linking to fix protein-DNA interactions. Subsequently, nuclei were isolated and chromatin was fragmented into 200–500 bp by sonication. Flag antibody was used for specific immunoprecipitation of BpTCP20-DNA complexes, with Input and negative controls set to ensure result reliability. After reverse cross-linking and purification, the precipitated DNA was used for ChIP-seq library construction and high-throughput sequencing. Finally, bioinformatics analyses (including sequence alignment, Peak calling, annotation, and Motif analysis) were performed to identify the precise binding sites of BpTCP20 genome-wide.
Northeast Forestry University