Targeting Induced Local Lesions IN Genomes (TILLING) is a reverse genetics method enabling the identification of individuals carrying chemically induced allelic variants at a gene of interest, within a mutagenized population. TILLING was first described in Arabidopsis (McCallum et al., 2000) and Drosophila (Bentley et al., 2000), and then extended to several plant species including maize (Till et al., 2004b), wheat (Slade et al., 2005), rice (Suzuki et al., 2008; Till et al., 2007), and others. It can be implemented in any species where a population of mutant individuals can be generated starting from a homogeneous line. Additionally, a version of TILLING has been described to help in discovery of polymorphisms in natural populations or in cultivars collections (EcoTILLING; Comai et al., 2004). Commonly, TILLING a gene implies three consecutive steps, namely (i) production or availability of a mutagenized population, (ii) PCR amplification of the target gene in a pool of individual genomic DNA samples, and (iii) detection of mutant alleles and individuals carrying them by means of a biochemical essay suitable for nucleotide polymorphism (e.g. SNP) identification. Most TILLING resources have been developed using chemical mutagenesis which mostly induces single-point mutations. This provides the investigator with a very informative range of allelic effects, from effective protein knockouts produced by stop-gain mutations to hypomorphic alleles caused by missense (i.e. amino acid substitution) mutations. Such different functional alleles are not usually available within populations generated with other types of mutational agents such as insertional mutagenesis (e.g. T-DNA, transposons) or irradiation (e.g. gamma-ray, fast-neutrons), all of which mostly generate knockouts. One additional positive aspect of TILLING is that it does not require any prior genomic knowledge (except for the target gene sequence, which must be known).

Advances in molecular breeding techniques for barley: targeted induced local lesions in genomes (TILLING)

Rosignoli, Serena;Salvi, Silvio
2020

Abstract

Targeting Induced Local Lesions IN Genomes (TILLING) is a reverse genetics method enabling the identification of individuals carrying chemically induced allelic variants at a gene of interest, within a mutagenized population. TILLING was first described in Arabidopsis (McCallum et al., 2000) and Drosophila (Bentley et al., 2000), and then extended to several plant species including maize (Till et al., 2004b), wheat (Slade et al., 2005), rice (Suzuki et al., 2008; Till et al., 2007), and others. It can be implemented in any species where a population of mutant individuals can be generated starting from a homogeneous line. Additionally, a version of TILLING has been described to help in discovery of polymorphisms in natural populations or in cultivars collections (EcoTILLING; Comai et al., 2004). Commonly, TILLING a gene implies three consecutive steps, namely (i) production or availability of a mutagenized population, (ii) PCR amplification of the target gene in a pool of individual genomic DNA samples, and (iii) detection of mutant alleles and individuals carrying them by means of a biochemical essay suitable for nucleotide polymorphism (e.g. SNP) identification. Most TILLING resources have been developed using chemical mutagenesis which mostly induces single-point mutations. This provides the investigator with a very informative range of allelic effects, from effective protein knockouts produced by stop-gain mutations to hypomorphic alleles caused by missense (i.e. amino acid substitution) mutations. Such different functional alleles are not usually available within populations generated with other types of mutational agents such as insertional mutagenesis (e.g. T-DNA, transposons) or irradiation (e.g. gamma-ray, fast-neutrons), all of which mostly generate knockouts. One additional positive aspect of TILLING is that it does not require any prior genomic knowledge (except for the target gene sequence, which must be known).
Achieving sustainable cultivation of barley
203
224
Rosignoli, Serena; Salvi, Silvio
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11585/743151
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