GeneCalling
In the field of genomics, GeneCalling is an open-platform mRNA transcriptional profiling technique.{{cite journal|last1=Kirst|first1=M. E.|title=Identification and Characterization of Endoplasmic Reticulum-Associated Degradation Proteins Differentially Affected by Endoplasmic Reticulum Stress|journal=Plant Physiology |date=1 May 2005|volume=138|issue=1|pages=218–231|doi=10.1104/pp.105.060087|pmc=1104177|pmid=15849299}} The GeneCalling protocol measures levels of cDNA, which are correlated with gene expression levels of specific transcripts. Differences between gene expression in healthy tissues and disease or drug responsive tissues are examined and compared in this technology.{{cite journal|last1=Green|first1=Cynthia D.|last2=Simons|first2=Jan Fredrik|last3=Taillon|first3=Bruce E.|last4=Lewin|first4=David A.|title=Open systems: panoramic views of gene expression|journal=Journal of Immunological Methods|date=April 2001|volume=250|issue=1–2|pages=67–79|doi=10.1016/S0022-1759(01)00306-4|pmid=11251222}} The technique has been applied to the study of human tissuesGene Expression Analysis by Transcript Profiling Coupled to a Gene Database Query. Nature Biotechnology 17, 798-803: 1999 and plant tissues.Expression Profiling of the Maize Flavonoid Pathway Genes Controlled by Estradiol-Inducible Transcription Factors CRC and P. The Plant Cell 12, 65-79: 2000
Method
In the GeneCalling protocol, mRNAs are first isolated from a given sample and processed into fragments for analysis. This usually involves the synthesis and subdivision of double-stranded cDNAs from polyA RNA. Distinct sets of restriction enzymes can then be used to digest sets of the divided cDNAs and resulting fragments ligated to labelled adapters to be amplified by PCR. PCR products are then purified and subjected to gel electrophoresis on a mounted platform employing stationary laser excitation and a multi-colour charge-coupled device imaging system.{{cite journal|last1=Klein|first1=S|last2=de Fougerolles|first2=AR|last3=Blaikie|first3=P|last4=Khan|first4=L|last5=Pepe|first5=A|last6=Green|first6=CD|last7=Koteliansky|first7=V|last8=Giancotti|first8=FG|title=Alpha 5 beta 1 integrin activates an NF-kappa B-dependent program of gene expression important for angiogenesis and inflammation.|journal=Molecular and Cellular Biology|date=August 2002|volume=22|issue=16|pages=5912–22|pmid=12138201|pmc=133962|doi=10.1128/mcb.22.16.5912-5922.2002}} A fluorescent label at the 5' end of one of the PCR primers allows for visualization of the PCR fragments, and the cDNAs are subjected to several isolated and identical restriction digests to generate a merged profile based on peak height and variance.{{cite journal | last1 = Shimkets | first1 = R.A. | display-authors = etal | year = 1999 | title = Gene Expression Analysis by Transcript Profiling Coupled to a Gene Database Query | journal = Nat. Biotechnol. | volume = 17 | issue = 8| pages = 798–803 | doi = 10.1038/11743 | pmid = 10429247 | s2cid = 25463457 }} The merged digestion profiles from the cDNA preparations are then compared to locate differentially expressed fragments (such as between normal tissue and diseased or drug responsive tissue); these profiles are compared by means of various internet-ready databases such as GeneScape.{{cite book|last1=Grotewold|first1=Erich|title=Plant Functional Genomics|date=2003|publisher=Springer Science & Business Media|isbn=9781592594139|page=386|url=https://books.google.com/books?id=E3A_aXGjGS8C&q=Genecalling+genescape&pg=PA386|access-date=17 October 2016|language=en}}