Galbraith, D. (2012). Flow Cytometry and Cell Sorting: the Next Generation. Methods.
Grebenok, R. J., Pierson, E., Lambert, G. M., Gong, F., Afonso, C. L., Haldeman-Cahill, R., Carrington, J. C., & Galbraith, D. W. (1997). Green-fluorescent protein fusions for efficient characterization of nuclear targeting. Plant Journal, 11(3), 573-586.
PMID: 9107043;Abstract:
The green-fluorescent protein (GFP) from Aequorea victoria has been shown to be a convenient and flexible reporter molecule within a variety of eukaryotic systems, including higher plants. It is particularly suited for applications in vivo, since the mechanism of fluorophore formation involves an intramolecular autoxidation and does not require exogenous co-factors. Unlike standard histochemical procedures of fixation and staining required for analysis of the cellular or tissue-specific expression of other popular reporter molecules, such as the β-glucuronidase (GUS) marker, analysis of GFP can be done in living cells with no specific pretreatments. This implies that GFP might also be particularly suited for studies of intracellular protein targeting. In this paper, the use of GUS is compared with that of GFP for the analysis of nuclear targeting in tobacco. A novel oligopeptide motif from a tobacco protein is described which confers nuclear localization of GUS. The use of this oligopeptide and two from potyviral proteins to target GFP to the nucleus is examined. An essential modification of GFP is described, which specifically increases its molecular weight to eliminate its passive penetration into the nucleus. Three examples of the targeting of these enlarged GFP molecules to the nucleus are illustrated. GFP, in combination with confocal microscopy, offers significant advantages over traditional methods of studying nuclear targeting.
Galbraith, D. (2011). The grand challenges in enabling data-intensive biological research. Frontiers in Genetics, 2(26).
doi:10.3389fgene.2011. 00026
Weng, Z., Barthelson, R., Gowda, S., Hilf, M. E., Dawson, W. O., Galbraith, D. W., & Xiong, Z. (2007). Persistent infection and promiscuous recombination of multiple genotypes of an RNA virus within a single host generate extensive diversity. PLoS ONE, 2(9).
PMID: 17878952;PMCID: PMC1975466;Abstract:
Recombination and reassortment of viral genomes are major processes contributing to the creation of new, emerging viruses. These processes are especially significant in long-term persistent infections where multiple viral genotypes co-replicate in a single host, generating abundant genotypic variants, some of which may possess novel host-colonizing and pathogenicity traits. In some plants, successive vegetative propagation of infected tissues and introduction of new genotypes of a virus by vector transmission allows for viral, populations to increase in complexity for hundreds of years allowing co-replication and subsequent recombination of the multiple viral genotypes. Using a resequencing microarray, we examined a persistent infection by a Citrus tristeza virus (CTV) complex in citrus, a vegetatively propagated, globally important fruit crop, and found that the complex comprised three major and a number of minor genotypes. Subsequent deep sequencing analysis of the viral population confirmed the presence of the three major CTV genotypes and, in addition, revealed that the minor genotypes consisted of an extraordinarily large number of genetic variants generated by promiscuous recombination between the major genotypes. Further analysis provided evidence that some of the recombinants underwent subsequent divergence, further increasing the genotypic complexity. These data demonstrate that persistent infection of multiple viral genotypes within a host organism is sufficient to drive the large-scale production of viral genetic variants that may evolve into new and emerging viruses. © 2007 Weng et al.
Rangwala, S. H., Elumalai, R., Vanier, C., Ozkan, H., Galbraith, D. W., & Richards, E. J. (2006). Meiotically stable natural epialleles of Sadhu, a novel arabidopsis retroposon. PLoS Genetics, 2(3), 0270-0281.
Abstract:
Epigenetic variation is a potential source of genomic and phenotypic variation among different individuals in a population, and among different varieties within a species. We used a two-tiered approach to identify naturally occurring epigenetic alleles in the flowering plant Arabidopsis: a primary screen for transcript level polymorphisms among three strains (Col, Cvi, Ler), followed by a secondary screen for epigenetic alleles. Here, we describe the identification of stable, meiotically transmissible epigenetic alleles that correspond to one member of a previously uncharacterized non-LTR retroposon family, which we have designated Sadhu. The pericentromeric At2g10410 element is highly expressed in strain Col, but silenced in Ler and 18 other strains surveyed. Transcription of this locus is inversely correlated with cytosine methylation and both the expression and DNA methylation states map in a Mendelian manner to stable cis-acting variation. The silent Ler allele can be converted by the epigenetic modifier mutation ddm1 to a meiotically stable expressing allele with an identical primary nucleotide sequence, demonstrating that the variation responsible for transcript level polymorphism among Arabidopsis strains is epigenetic. We extended our characterization of the Sadhu family members and show that different elements are subject to both genetic and epigenetic variation in natural populations. These findings support the view that an important component of natural variation in retroelements is epigenetic. Copyright: © 2006 Rangwala et al.
