Tilling (molecular biology)

TILLING ("Targeting Induced Local Lesions in Genomes") is a method in molecular biology that allows directed identification of mutations in a specific gene.

The method combines a standard technique, mutagenesis with a chemical mutagen such as Ethyl methanesulfonate (EMS) with a sensitive DNA screening-technique that identifies single base mutations (also called point mutations) in a target gene. The first paper describing TILLING used dHPLC to identify mutations (McCallum et al., 2000a) . The method was made more high throughput by using the restriction enzyme Cel-I combined with a gel based system to identify mutations (Colbert et al.,2001). Other methods of mutation detection, such as resequencing DNA, have been combined for TILLING.

TILLING was introduced in 2000, using the model plant Arabidopsis. TILLING has since been used as a reverse genetics method in other organisms such as zebrafish, corn, wheat, rice, soybean, tomato and lettuce.

References

McCallum CM, Comai L, Greene EA, Henikoff S. Targeted screening for induced mutations. Nat Biotechnol. 2000 Apr;18(4):455-7.

McCallum CM, Comai L, Greene EA, Henikoff S. Targeting induced local lesions IN genomes (TILLING) for plant functional genomics. Plant Physiol. 2000 Jun;123(2):439-42.

Colbert T, Till BJ, Tompa R, Reynolds S, Steine MN, Yeung AT, McCallum CM, Comai L, Henikoff S. High-throughput screening for induced point mutations. Plant Physiol. 2001 Jun;126(2):480-4.

Draper BW, McCallum CM, Stout JL, Slade AJ, Moens CB. A high-throughput method for identifying N-ethyl-N-nitrosourea (ENU)-induced point mutations in zebrafish. Methods Cell Biol. 2004;77:91-112.

Slade AJ, Fuerstenberg SI, Loeffler D, Steine MN, Facciotti D. A reverse genetic, nontransgenic approach to wheat crop improvement by TILLING. Nat Biotechnol. 2005 Jan;23(1):75-81.

External links

* [http://tilling.fhcrc.org:9366/files/Welcome_to_ATP.html The Arabidopsis Tilling project]
* [http://www.licor.com/bio/genomics/Tilling/TillingMain.jsp Introduction to TILLING]

reverse genetics