Editing Reporter gene (section)

==Gene expression assays==
Reporter genes can be used to assay for the expression of a gene of interest that is normally difficult to quantitatively assay.<ref name=":5" /> Reporter genes can produce a protein that has little obvious or immediate effect on the cell culture or organism. They are ideally not present in the native genome to be able to isolate reporter gene expression as a result of the gene of interest's expression.<ref name=":5" /><ref>Archived at [https://ghostarchive.org/varchive/youtube/20211211/PD_6JU3NayE Ghostarchive]{{cbignore}} and the [https://web.archive.org/web/20200614175119/https://www.youtube.com/watch?v=PD_6JU3NayE&gl=US&hl=en Wayback Machine]{{cbignore}}: {{Cite web|url=https://www.youtube.com/watch?v=PD_6JU3NayE|title=Introduction to Reporter Gene Assays|last=Promega Corporation|first=Promega Corporation|date=October 22, 2014|website=YouTube|access-date=March 21, 2020}}{{cbignore}}</ref>

To activate reporter genes, they can be expressed [[Gene expression|constitutively]], where they are directly attached to the gene of interest to create a [[gene fusion]].<ref>{{Cite book|last1=de Jong|first1=Hidde|last2=Geiselmann|first2=Johannes|title=Hybrid Systems Biology |chapter=Fluorescent Reporter Genes and the Analysis of Bacterial Regulatory Networks |date=2015|editor-last=Maler|editor-first=Oded|editor2-last=Halász|editor2-first=Ádám|editor3-last=Dang|editor3-first=Thao|editor4-last=Piazza|editor4-first=Carla|volume=7699|series=Lecture Notes in Computer Science|language=en|publisher=Springer International Publishing|pages=27–50|doi=10.1007/978-3-319-27656-4_2|isbn=978-3-319-27656-4}}</ref> This method is an example of using [[Cis-regulatory element|''cis''-acting]] elements where the two genes are under the same promoter elements and are [[Transcription (genetics)|transcribed]] into a single [[messenger RNA]] molecule. The [[mRNA]] is then [[Translation (biology)|translated]] into protein. It is important that both proteins be able to properly [[protein folding|fold]] into their active conformations and interact with their substrates despite being fused. In building the DNA construct, a segment of DNA coding for a flexible polypeptide linker region is usually included so that the reporter and the gene product will only minimally interfere with one another.<ref>{{Cite journal|last1=Spector|first1=David L.|last2=Goldman|first2=Robert D.|date=2006-12-01|title=Constructing and Expressing GFP Fusion Proteins|journal=Cold Spring Harbor Protocols|volume=2006|issue=7|pages=pdb.prot4649|doi=10.1101/pdb.prot4649|pmid=22484672}}</ref><ref>{{Cite journal|last1=Chen|first1=Xiaoying|last2=Zaro|first2=Jennica|last3=Shen|first3=Wei-Chiang|date=2013-10-15|title=Fusion Protein Linkers: Property, Design and Functionality|journal=Advanced Drug Delivery Reviews|volume=65|issue=10|pages=1357–1369|doi=10.1016/j.addr.2012.09.039|issn=0169-409X|pmc=3726540|pmid=23026637}}</ref> Reporter genes can also be expressed by [[Gene expression|induction]] during growth. In these cases, [[Trans-acting|''trans''-acting]] elements, such as [[transcription factor]]s are used to express the reporter gene.<ref>{{Citation|last1=Hanko|first1=Erik K. R.|chapter=Chapter Nine - Design, cloning and characterization of transcription factor-based inducible gene expression systems|date=2019-01-01|url=http://www.sciencedirect.com/science/article/pii/S0076687919300370|series=Methods in Enzymology|volume=621|pages=153–169|editor-last=Shukla|editor-first=Arun K.|title=Chemical and Synthetic Biology Approaches To Understand Cellular Functions - Part A|publisher=Academic Press|access-date=2019-12-16|last2=Minton|first2=Nigel P.|last3=Malys|first3=Naglis|doi=10.1016/bs.mie.2019.02.018|pmid=31128776|isbn=978-0-12-818117-1 |s2cid=91744525|url-access=subscription}}</ref><ref>{{Cite journal|last1=Kallunki|first1=Tuula|last2=Barisic|first2=Marin|last3=Jäättelä|first3=Marja|last4=Liu|first4=Bin|date=2019-07-30|title=How to Choose the Right Inducible Gene Expression System for Mammalian Studies?|journal=Cells|volume=8|issue=8|page=796|doi=10.3390/cells8080796|issn=2073-4409|pmc=6721553|pmid=31366153|doi-access=free}}</ref>

Reporter gene assay have been increasingly used in [[High-throughput screening|high throughput screening]] (HTS) to identify small molecule inhibitors and activators of protein targets and pathways for [[drug discovery]] and [[chemical biology]].  Because the reporter enzymes themselves (e.g. firefly [[luciferase]]) can be direct targets of small molecules and confound the interpretation of HTS data, novel coincidence reporter designs incorporating artifact suppression have been developed.<ref>{{cite journal | title = A coincidence reporter-gene system for high throughput screening. |author1 = Cheng, K.C.| author2 = Inglese, J. | journal = Nature Methods|date = 2012 |volume = 9|issue = 10|page = 937|pmid = 23018994 | doi = 10.1038/nmeth.2170 | pmc=4970863}}</ref><ref>{{cite journal | title = Chemogenomic profiling of endogenous PARK2 expression using a genome-edited coincidence reporter. |author1 = Hasson, S.A.|author2 = Fogel, A.I.|author3 = Wang, C.| author4 = MacArthur, R.|author5 = Guha, R.|author6 = Heman-Ackahc, S.|author7 = Martin, S.| author8 = Youle, R.J.|author9 = Inglese, J. | journal = ACS Chem. Biol.|date = 2015 |volume = 10|issue = 5|pages = 1188–1197|pmid = 25689131| doi = 10.1021/cb5010417| pmc=9927027 | s2cid=20139739 }}</ref>