Editing Functional genomics (section)

===At the protein level===

====Yeast two-hybrid system====
{{Main|Two-hybrid screening}}
A yeast [[two-hybrid screening]] (Y2H) tests a "bait" protein against many potential interacting proteins ("prey") to identify physical protein–protein interactions. This system is based on a transcription factor, originally GAL4,<ref name=pmid2547163>{{cite journal | vauthors = Fields S, Song O | title = A novel genetic system to detect protein-protein interactions | journal = Nature | volume = 340 | issue = 6230 | pages = 245–6 | date = July 1989 | pmid = 2547163 | doi = 10.1038/340245a0 | bibcode = 1989Natur.340..245F | s2cid = 4320733 }}</ref> whose separate DNA-binding and transcription activation domains are both required in order for the protein to cause transcription of a reporter gene. In a Y2H screen, the "bait" protein is fused to the binding domain of GAL4, and a library of potential "prey" (interacting) proteins is recombinantly expressed in a vector with the activation domain. In vivo interaction of bait and prey proteins in a yeast cell brings the activation and binding domains of GAL4 close enough together to result in expression of a [[reporter gene]]. It is also possible to systematically test a library of bait proteins against a library of prey proteins to identify all possible interactions in a cell.

====MS and AP/MS====
{{Main|Protein mass spectrometry|Affinity purification}}
[[Mass spectrometry]] (MS) can identify proteins and their relative levels, hence it can be used to study protein expression. When used in combination with [[affinity purification]], [[mass spectrometry]] (AP/MS) can be used to study protein complexes, that is, which proteins interact with one another in complexes and in which ratios. In order to purify protein complexes, usually a "bait" protein is tagged with a specific protein or peptide that can be used to pull out the complex from a complex mix. The purification is usually done using an antibody or a compound that binds to the fusion part. The proteins are then digested into short [[peptide]] fragments and mass spectrometry is used to identify the proteins based on the mass-to-charge ratios of those fragments.

====Deep mutational scanning====
In deep mutational scanning, every possible amino acid change in a given protein is first synthesized.<ref>{{cite journal |last1=Araya |first1=Carlos |last2=Fowler |first2=Douglas |title=Deep mutational scanning: assessing protein function on a massive scale |journal=Trends in Biotechnology |date=September 29, 2011 |volume=29 |issue=9 |pages=435–442 |doi=10.1016/j.tibtech.2011.04.003 |pmid=21561674|pmc=3159719 }}</ref> The activity of each of these protein variants is assayed in parallel using barcodes for each variant.<ref>{{cite journal | vauthors = Penn WD, McKee AG, Kuntz CP, Woods H, Nash V, Gruenhagen TC, Roushar FJ, Chandak M, Hemmerich C, Rusch DB, Meiler J, Schlebach JP| title = Probing biophysical sequence constraints within the transmembrane domains of rhodopsin by deep mutational scanning| journal = Sci Adv | volume =  6 | issue = 10 | pages = eaay7505| date = March 2020 | pmid = 32181350 | doi = 10.1126/sciadv.aay7505| pmc = 7056298 | bibcode = 2020SciA....6.7505P}}</ref> By comparing the activity to the wild-type protein, the effect of each mutation is identified. While it is possible to assay every possible single amino-acid change due to combinatorics two or more concurrent mutations are hard to test. Deep mutational scanning experiments have also been used to infer protein structure and protein-protein interactions.<ref>{{cite journal |last1=Rollins |first1=N.J. |last2=Brock |first2=K.P. |last3=Poelwijk |first3=F.J |last4=Marks |first4=Debora |title=Inferring protein 3D structure from deep mutation scans |journal=Nature Genetics |date=2019 |volume=51 |issue=7 |pages=1170–1176 |doi=10.1038/s41588-019-0432-9 |pmid=31209393 |pmc=7295002 }}</ref> Deep Mutational Scanning is an example of a multiplexed assays of variant effect (MAVEs), a family of methods that involve mutagenesis of a DNA-encoded protein or regulatory element followed by a multiplexed assay for some aspect of function. MAVEs enable the generation of ‘variant effect maps’ characterizing aspects of the function of every possible single nucleotide change in a gene or functional element of interest. <ref>{{cite journal |last1=Fowler |first1=DM |last2=Adams |first2=DJ |last3=Gloyn |first3=AL |last4=Starita |first4=Lea |title=An Atlas of Variant Effects to understand the genome at nucleotide resolution |journal=Genome Biology |date=2023 |volume=24 |issue=1 |page=147 |doi=10.1186/s13059-023-02986-x |doi-access=free |pmid=37394429|pmc=10316620 }}</ref>