Anderson, United States
Anderson, United States

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Li T.,Development and Cell Biology | Huang S.,Development and Cell Biology | Zhao X.,Laurence ker Center For Bioinformatics And Biological Statistics | Wright D.A.,Development and Cell Biology | And 4 more authors.
Nucleic Acids Research | Year: 2011

Recent studies indicate that the DNA recognition domain of transcription activator-like (TAL) effectors can be combined with the nuclease domain of FokI restriction enzyme to produce TAL effector nucleases (TALENs) that, in pairs, bind adjacent DNA target sites and produce double-strand breaks between the target sequences, stimulating non-homologous end-joining and homologous recombination. Here, we exploit the four prevalent TAL repeats and their DNA recognition cipher to develop a 'modular assembly' method for rapid production of designer TALENs (dTALENs) that recognize unique DNA sequence up to 23 bases in any gene. We have used this approach to engineer 10 dTALENs to target specific loci in native yeast chromosomal genes. All dTALENs produced high rates of site-specific gene disruptions and created strains with expected mutant phenotypes. Moreover, dTALENs stimulated high rates (up to 34) of gene replacement by homologous recombination. Finally, dTALENs caused no detectable cytotoxicity and minimal levels of undesired genetic mutations in the treated yeast strains. These studies expand the realm of verified TALEN activity from cultured human cells to an intact eukaryotic organism and suggest that low-cost, highly dependable dTALENs can assume a significant role for gene modifications of value in human and animal health, agriculture and industry. © 2011 The Author(s).

Zuo T.,Development and Cell Biology | Zhang J.,Development and Cell Biology | Lithio A.,Iowa State University | Dash S.,Iowa State University | And 4 more authors.
Genetics | Year: 2016

Copy-number alterations are widespread in animal and plant genomes, but their immediate impact on gene expression is still unclear. In animals, copy-number alterations usually exhibit dosage effects, except for sex chromosomes which tend to be dosage compensated. In plants, genes within small duplications (<100 kb) often exhibit dosage-dependent expression, whereas large duplications (>50 Mb) are more often dosage compensated. However, little or nothing is known about expression in moderately-sized (1- 50 Mb) segmental duplications, and about the response of small RNAs to dosage change. Here, we compared maize (Zea mays) plants with two, three, and four doses of a 14.6-Mb segment of chromosome 1 that contains ~300 genes. Plants containing the duplicated segment exhibit dosage-dependent effects on ear length and flowering time. Transcriptome analyses using GeneChip and RNAsequencing methods indicate that most expressed genes and unique small RNAs within the duplicated segments exhibit dosagedependent transcript levels. We conclude that dosage effect is the predominant regulatory response for both genes and unique small RNA transcripts in the segmental dosage series we tested. To our knowledge this is the first analysis of small RNA expression in plant gene dosage variants. Because segmental duplications comprise a significant proportion of eukaryotic genomes, these findings provide important new insight into the regulation of genes and small RNAs in response to dosage changes. © 2016 by the Genetics Society of America.

PubMed | Development and Cell Biology, Iowa State University and University of Wisconsin - Stevens Point
Type: Journal Article | Journal: Plants (Basel, Switzerland) | Year: 2016

In soybean, genic male sterility can be utilized as a tool to develop hybrid seed. Several male-sterile, female-fertile mutants have been identified in soybean. The male-sterile, female-fertile ms5 mutant was selected after fast neutron irradiation. Male-sterility due to ms5 was associated with the stay-green cotyledon color mutation. The cotyledon color trait in soybean is controlled by two loci, D and D. Association between cotyledon color and male-sterility can be instrumental in early phenotypic selection of sterility for hybrid seed production. The use of such selection methods saves time, money, and space, as fewer seeds need to be planted and screened for sterility. The objectives of this study were to compare anther development between male-fertile and male-sterile plants, to investigate the possible linkages among the Ms5, D and D loci, and to determine if any of the d or d mutations can be applied in hybrid seed production. The cytological analysis during anther development displayed optically clear, disintegrating microspores and enlarged, engorged pollen in the male-sterile, female-fertile ms5ms5 plants, a common characteristic of male-sterile mutants. The D locus was mapped to molecular linkage group (MLG) D1a and was flanked by Satt408 and BARCSOYSSR_01_1622. The ms5 and D loci were mapped to MLG B1 with a genetic distance ~12.8 cM between them. These results suggest that use of the d mutant in the selection of male-sterile line may attenuate the cost hybrid seed production in soybean.

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