A functional genomics resource for <i>Brassica napus</i>: development of an EMS mutagenized population and discovery of <i>FAE1</i> point mutations by TILLING

Wang, Nian; Wang, Yajie; Tian, Fang; King, Graham J.; Zhang, Chunyu; Long, Yan; Shi, Lei; Meng, Jianghong

doi:10.1111/j.1469-8137.2008.02619.x

Cited by 140 publications

(96 citation statements)

References 49 publications

(83 reference statements)

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“…The highest mutation densities were obtained in polyploids ( Fig. 1), hexaploid wheat (Triticum aestivum; Slade et al, 2005;Uauy et al, 2009), and tetraploid canola (Brassica napus; Wang et al, 2008;Harloff et al, 2012), displaying the highest number at approximately 40 mutations Mb -1 of diploid DNA (i.e. for each of the ancestral genomes of the polyploid).…”

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confidence: 99%

Production of a High-Efficiency TILLING Population through Polyploidization

et al. 2013

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Targeting Induced Local Lesions in Genomes (TILLING) provides a nontransgenic method for reverse genetics that is widely applicable, even in species where other functional resources are missing or expensive to build. The efficiency of TILLING, however, is greatly facilitated by high mutation density. Species vary in the number of mutations induced by comparable mutagenic treatments, suggesting that genetic background may affect the response. Allopolyploid species have often yielded higher mutation density than diploids. To examine the effect of ploidy, we autotetraploidized the Arabidopsis (Arabidopsis thaliana) ecotype Columbia, whose diploid has been used for TILLING extensively, and mutagenized it with 50 mM ethylmethane sulfonate. While the same treatment sterilized diploid Columbia, the tetraploid M1 plants produced good seed. To determine the mutation density, we searched 528 individuals for induced mutations in 15 genes for which few or no knockout alleles were previously available. We constructed tridimensional pools from the genomic DNA of M2 plants, amplified target DNA, and subjected them to Illumina sequencing. The results were analyzed with an improved version of the mutation detection software CAMBa that accepts any pooling scheme. This small population provided a rich resource with approximately 25 mutations per queried 1.5-kb fragment, including on average four severe missense and 1.3 truncation mutations. The overall mutation density of 19.4 mutations Mb -1 is 4 times that achieved in the corresponding diploid accession, indicating that genomic redundancy engenders tolerance to high mutation density. Polyploidization of diploids will allow the production of small populations, such as less than 2,000, that provide allelic series from knockout to mild loss of function for virtually all genes.

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confidence: 99%

Production of a High-Efficiency TILLING Population through Polyploidization

et al. 2013

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“…The mutation frequency in this population is, therefore, approximately 1 in 447 kb ((12,525 £ 893)/25). The frequency and spec trum of mutations in this species was not substantially diVerent from that observed in TILLING populations of other diploid species such as barley (1 mutation per 374 kb: Talamè et al 2008), Arabidopsis (1 mutation per 300 kb: Greene et al 2003) and rice (1 mutation per 300 kb: Till et al 2007), but lower than that seem in polyploids such as wheat (1 mutation per 24 kb: Slade et al 2005) or Brassica napus (1 mutation per 41.5 kb: Wang et al 2008). This is as expected, since genetic redundancy in polyploids appears to allow higher mutational loads in these species.…”

Section: Population Constructionmentioning

confidence: 96%

Forward and reverse genetics of rapid-cycling Brassica oleracea

et al. 2009

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Seeds of rapid-cycling Brassica oleracea were mutagenized with the chemical mutagen, ethylmethane sul fonate. The reverse genetics technique, TILLING, was used on a sample population of 1,000 plants, to determine the mutation proWle. The spectrum and frequency of mutations induced by ethylmethane sulfonate was similar to that seen in other diploid species such as Arabidopsis thaliana. These data indicate that the mutagenesis was eVective and demonstrate that TILLING represents an eYcient reverse genetic technique in B. oleracea that will become more valuable as increasing genomic sequence data become available for this species. The extensive duplication in the B. oleracea genome is believed to result in the genetic redundancy that has been important for the evolution of morphological diversity seen in today's B. oleracea crops (broccoli, Brussels sprouts, cauliXower, cabbage, kale and kohlrabi). However, our forward genetic screens identiWed 120 mutants in which some aspect of development was

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“…TILLING involves induction of mutations in the plant genome using classical mutagenesis approaches followed by traditional or high throughput deep sequencing to identify the mutations in the gene of interest [77][78][79]. This technique has been used in allele discovery in different plant species [80][81][82][83]. EcoTILLING, which is an adaptation of the TILLING, is used in detecting rare single nucleotide polymorphism (SNPs) or small INDELs in target genes in natural populations [84].…”

Section: Tilling and Ecotillingmentioning

confidence: 99%

Recent Biotechnological Advances in the Improvement of Cassava

Fondong¹,

Rey²

2018

Cassava

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Cassava (Manihot esculenta Crantz) is the fourth most important source of carbohydrates for human consumption in the tropics and thus occupies a uniquely important position as a food security crop for smallholder farmers. Consequently, cassava improvement is of high priority to most national agricultural research institutions in the tropics. With advances in functional genomics and genome editing approaches in this post genomics era, there are unprecedented opportunities and potential to accelerate the improvement of this important crop. These new technologies will need to be directed toward addressing major cassava production constraints, notably virus resistance, protein content, tolerance to drought and reduction of hydrogen cyanide content. Here, we discuss the important role novel functional genomics and genome editing technologies have and will continue to play in cassava improvement efforts. These approaches, including artificial miRNA (amiRNA), trans-acting small interfering RNA (tasiRNA), clustered regularly interspaced short palindromic repeat (CRISPR)-associated protein 9 (Cas9), and Targeting Induced Local Lesions IN Genomes (TILLING), have been shown to be effective in addressing major crop production constraints. In addition to reviewing specific applications of these technologies in cassava improvement, this chapter discusses specific examples being deployed in the amelioration of cassava or of other crops that could be applied to cassava in future.

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A functional genomics resource for Brassica napus: development of an EMS mutagenized population and discovery of FAE1 point mutations by TILLING

Cited by 140 publications

References 49 publications

Production of a High-Efficiency TILLING Population through Polyploidization

Production of a High-Efficiency TILLING Population through Polyploidization

Forward and reverse genetics of rapid-cycling Brassica oleracea

Recent Biotechnological Advances in the Improvement of Cassava

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