SOSHI-seq: a high-throughput screening assay to test the functionality of putative response elements for nuclear hormone receptors.

We present a cheap, rapid, and versatile assay called SOSHI-seq (Screening Of Self-transcribed Hormone Inducible response elements coupled to sequencing). The protocol is an adaptation of the STARR-seq method, which allows testing the capacity of thousands of synthetic DNA sequences to act as hormone-response elements. Using the nuclear receptors of thyroid hormone as an example, we show that SOSHI-seq is a suitable complement to ChIP-Seq analysis to identify at genome-wide scale the functional response elements occupied by nuclear receptors in chromatin. Overall design: SOSHI-seq: a high-throughput screening assay to test the functionality of putative response elements for nuclear hormone receptors. SOSHI-seq: a high-throughput screening assay to test the functionality of putative response elements for nuclear hormone receptors. Banks are prepared with 1000 inserts in a plasmid vector in which sequences able to bind a nuclear receptor are inserted. Plasmid banks are transfected in HEK293 cells and RNA is extracted. Amplicons are prepared, corresponding to the variable part of the plasmid transcript. Amplicons are used to prepare sequencing libraries. Samples differ by : - the plasmid bank that is transfected (bank1 or bank2) - the type of nuclear receptor that is encoded by an additionnal plasmid that is cotransfected with the bank (TRa1, TRß1 or LXRa) - the presence or absence of agonist in the culture medium (thyroid hormone , or T3, for TRa1, TRß1, T-compound for LXRa). Some of the experiments are duplicated.