JRD278Chip-SEQ protein technical service

CHIP-SEQ protein technical service brief introduction: chromatin immunocarpose: chromatin immune co-precipitation technology It is mainly used to verify and explore the direct interaction of proteins and DNA in vivo environments, intended to explore specific binding sequences of specific proteins in DNA, often applied in research transcriptional regulation and histone modification.

The experimental process is roughly: cross-linking-unconnected-ultrasound-immunoprecipitation-washing-DNA purification-PCR detection.

Specific operation:
Sample preparation: Cultured cells (if the sample is tissue, first cultivate primary cells); fixed, or directly live cells.

Cell Fixed: This step is to intend to crosslink the protein bonded to the DNA in a covalent form to avoid loss in the subsequent operating step. During the addition of formaldehyde, the cell culture medium was directly added to 10 minutes, and 2.5 M glycine decryplications were rapidly added for 5 minutes, PBS washing, and the cells were scraped.

Ultrasound Breaking: This step is to break the long chain DNA into 200-1000 bp DNA fragment. Using the equipment is an ultrasonic water bath, an EP tube is ultrasonic in a mixture of ice water, and the cell suspension has become clear and transparent. Centrifuge tap.

Immunoprecipitation: This step is to add specific antibodies and control IgG, and the magnetic beads (or Protein A / g) of the binding antibody. After dilution, the ultrasonic product was divided into two parts, and the specific antibody and control IgG were added, and the rotation fluid was added, and the magnetic bead wheel was added for 4 hours.

Washing: This step is to reduce non-specific binding. A total of 7 times was washed with a dilution buffer, a low salt solution, a high salt solution, a lithium chloride solution, and a TE buffer. Use the elution buffer eluting over the liquid at 65 ° C.

DNA purification: The eluted product was purified with a DNA purified kit to give 50 ul purified product for detecting CHIP results.

If you are interested in our Chip-SEQ, consult, we will warmly serve you.