A VISUAL AID FOR STATISTICIANS AND MOLECULAR BIOLOGISTS WORKING WITH MICROARRAY EXPERIMENTS

The use of microarrays to measure the expression of large numbers of genes simultaneously is increasing in agriculture research. Statisticians are expected to help biologists analyze these large data sets to identify biologically important genes that are differentially regulated in the samples under investigation. However, molecular biologists are often unfamiliar with the statistical methods used to analyze microarrays. Presented here are methods developed to graphically represent microarray data and various types of errors commonly associated with microarrays to help visualize sources of error. Two case studies were used. In case study one, genes differentially regulated when two corn lines, one resistant and one sensitive, were treated with Aspergillus flavus isolate NRRL 3357 or left untreated were investigated. Analyses and images showing 3 types of variation are shown. Genes were ranked according to fold change and re-ranked after adjusting for potential sources of error. In case two, cotton genes differentially regulated in 1-day-old fiber compared to whole ovules or older fibers were investigated. Data and sources of error were imaged as described for case one and genes with significant changes in gene expression were identified

Identification of Select Fumonisin Forming Fusarium Species Using PCR Applications of the Polyketide Synthase Gene and its Relationship to Fumonisin Production in vitro

A polymerase chain reaction (PCR) based diagnostic assay was used to develop markers for detection of Fusarium verticillioides (=F. moniliforme), a fumonisin producing fungus in maize tissues. Species-specific primers were designed based on sequence data from the polyketide synthase (PKS) gene (FUM1- previously FUM5) responsible for fumonisin production in fungi. Four sets of oligonucleotide primers were tested for their specificity using 24 strains of F. verticillioides, 10 F. proliferatum, and 12 of other Fusarium species. In addition, 13 species of other fungal genera, from four phyla, were tested as negative controls. Among the four sets, primer set B consistently amplified a 419-bp fragment from the DNA 96% of all F. verticillioides strains and 83% of F. proliferatum. All other fungi tested were negative using primer set B. A total of 38% of the F. verticillioides strains grown on a selective liquid medium produced fumonisin and 92% formed the toxin on standard rice medium. When fumonisin formed in culture, PCR assay using primer set B detected every strain of F. verticillioides, but only amplified 80% of F. proliferatum strains that produced the toxin. PCR detection was consistent at 100 pg/μl concentration of genomic DNA from 4 F. verticillioides strains, but varied at 10 pg/μl. Two duplicate greenhouse tests using artificially inoculated maize plants, had greater levels of F. verticillioides detected after re-evaluting using primer set B than from culturing of the tissues. The molecular protocols described in this study requires only 1 day for completion compared to approximately 10 days for cultural work and morphological determination. In conclusion, conventional PCR assay using primer set B provides a sensitive and accurate detection assay that can be used as a primary or secondary confirmation method for identification and occurrence of F. verticillioides within the maize tissues. However, studies using primer set B for fumonisin production determined by strains of F. verticillioides and F. proliferatum will require further verification

Hearing loss in a mouse model of Muenke syndrome

The heterozygous Pro250Arg substitution mutation in fibroblast growth factor receptor 3 (FGFR3), which increases ligand-dependent signalling, is the most common genetic cause of craniosynostosis in humans and defines Muenke syndrome. Since FGF signalling plays dosage-sensitive roles in the differentiation of the auditory sensory epithelium, we evaluated hearing in a large group of Muenke syndrome subjects, as well as in the corresponding mouse model (Fgfr3P244R). The Muenke syndrome cohort showed significant, but incompletely penetrant, predominantly low-frequency sensorineural hearing loss, and the Fgfr3P244R mice showed dominant, fully penetrant hearing loss that was more severe than that in Muenke syndrome individuals, but had the same pattern of relative high-frequency sparing. The mouse hearing loss correlated with an alteration in the fate of supporting cells (Deiters'-to-pillar cells) along the entire length of the cochlear duct, with the most extreme abnormalities found at the apical or low-frequency end. In addition, there was excess outer hair cell development in the apical region. We conclude that low-frequency sensorineural hearing loss is a characteristic feature of Muenke syndrome and that the genetically equivalent mouse provides an excellent model that could be useful in testing hearing loss therapies aimed at manipulating the levels of FGF signalling in the inner ear

Identification of Maize Genes Associated with Host Plant Resistance or Susceptibility to Aspergillus flavus Infection and Aflatoxin Accumulation

infection and aflatoxin accumulation. inoculation were compared in two resistant maize inbred lines (Mp313E and Mp04∶86) in contrast to two susceptible maize inbred lines (Va35 and B73) by microarray analysis. Principal component analysis (PCA) was used to find genes contributing to the larger variances associated with the resistant or susceptible maize inbred lines. The significance levels of gene expression were determined by using SAS and LIMMA programs. Fifty candidate genes were selected and further investigated by quantitative RT-PCR (qRT-PCR) in a time-course study on Mp313E and Va35. Sixteen of the candidate genes were found to be highly expressed in Mp313E and fifteen in Va35. Out of the 31 highly expressed genes, eight were mapped to seven previously identified quantitative trait locus (QTL) regions. A gene encoding glycine-rich RNA binding protein 2 was found to be associated with the host hypersensitivity and susceptibility in Va35. A nuclear pore complex protein YUP85-like gene was found to be involved in the host resistance in Mp313E. infection and aflatoxin accumulation. These findings will be important in identification of DNA markers for breeding maize lines resistant to aflatoxin accumulation

Lightning Talks and Research Grant Entire Program Video

This video contains multiple presentations from the Lightning Talks and the OLAC Research Grant presentation. Lightning Talks include: Rowena Griem: Locally Hosted Streaming Video Project at Yale University Library (time approx. 00:14:30), Kyla Jemison: Looking at Metadata in Digital Sheet Music Platforms (time approx. 00:24:20), Nicole Lewis: Online Exhibits: A Call for Cataloging Guidelines (time approx. 00:31:53), Amanda Mack: Challenges Cataloging Local News: The KTLA Tom Bradley Project (time approx. 00:39:41), and Adrian Williams: Cataloging Streaming Video at UK Libraries (time approx. 00:47:16). OLAC Research Grant presentation speakers are Michelle Urberg, Kelley McGrath and Morag Stewart, and the presentation is titled Acquisition, Cataloging and Discovery of Streaming Video. This presentation begins at time approx. 01:06:05

Primer sequences of candidate genes and the house keeping gene GAPDH in qRT-PCR study.

<p>Primer sequences of candidate genes and the house keeping gene GAPDH in qRT-PCR study.</p

2D plot showing gene expression levels over a period of 21 days after the fungal inoculation.

<p>These genes are highly expressed in Va35 than in Mp313E.</p