Biomedical subjects

Justen Andrews

Publications and source records attributed to Justen Andrews.

5 recordsLinked to original sources

PubMed2006

Automated liquid handling and high-throughput preparation of polymerase chain reaction-amplified DNA for microarray fabrication.

Genome-wide studies of gene expression and transcription factor-binding sites using DNA microarrays are leading to new systems level insights. The massively parallel nature of microarrays presents technical challenges: fabricating high-quality microarrays at the front end and data analysis and interpretation downstream. A principal challenge in fabricating microarrays is preparation of the DNA samples. This is particularly the case for polymerase chain reaction-amplified DNA samples. The challenge is to scale up efficiently to high-throughput preparation of tens of thousands of DNA samples while ensuring a uniform high quality. This chapter outlines strategic considerations, including automated liquid handling and workflow development to maximize efficiency, and quality control (QC) measures to ensure uniform quality. The protocols are presented with commentary to illustrate their logic and specific techniques. These principles and techniques are extensible to other high-throughput molecular biological applications.

Animals↗

PubMed2006

Troubleshooting microarray hybridizations.

Microarray experiments are being performed more widely than ever before, but even seasoned investigators can experience technical problems with hybridizations. This chapter provides guidelines for recognizing, rectifying, and avoiding common trouble areas. Specifically, it addresses frequent complications related to artifacts of printing, RNA sample preparation and quality, fluorophore labeling, hybridization conditions, and posthybridization washes. Emphasis is placed on investigating problems though a combination of appropriate controls and image analysis, where diagnostic plots of data quality are used to illustrate characteristics of acceptable and unsatisfactory hybridizations. This chapter also discusses resources available to microarray users hoping to improve the sensitivity and specificity of their experiments.

Oligonucleotide Array Sequence Analysis↗

PubMed2006

Microarray analysis reveals differential gene expression in hybrid sunflower species.

This paper describes the creation of a cDNA microarray for annual sunflowers and its use to elucidate patterns of gene expression in Helianthus annuus, Helianthus petiolaris, and the homoploid hybrid species Helianthus deserticola. The array comprises 3743 ESTs (expressed sequence tags) representing approximately 2897 unique genes. It has an average clone/EST identity rate of 91%, is applicable across species boundaries within the annual sunflowers, and shows patterns of gene expression that are highly reproducible according to real-time RT-PCR (reverse transcription-polymerase chain reaction) results. Overall, 12.8% of genes on the array showed statistically significant differential expression across the three species. Helianthus deserticola displayed transgressive, or extreme, expression for 58 genes, with roughly equal numbers exhibiting up- or down-regulation relative to both parental species. Transport-related proteins were strongly over-represented among the transgressively expressed genes, which makes functional sense given the extreme desert floor habitat of H. deserticola. The potential adaptive value of differential gene expression was evaluated for five genes in two populations of early generation (BC2) hybrids between the parental species grown in the H. deserticola habitat. One gene (a G protein-coupled receptor) had a significant association with fitness and maps close to a QTL controlling traits that may be adaptive in the desert habitat.

Base Sequence↗

PubMed2003

Paucity of genes on the Drosophila X chromosome showing male-biased expression.

Sex chromosomes are primary determinants of sexual dimorphism in many organisms. These chromosomes are thought to arise via the divergence of an ancestral autosome pair and are almost certainly influenced by differing selection in males and females. Exploring how sex chromosomes differ from autosomes is highly amenable to genomic analysis. We examined global gene expression in Drosophila melanogaster and report a dramatic underrepresentation of X-chromosome genes showing high relative expression in males. Using comparative genomics, we find that these same X-chromosome genes are exceptionally poorly conserved in the mosquito Anopheles gambiae. These data indicate that the X chromosome is a disfavored location for genes selectively expressed in males.

Animals↗

PubMed2002

Sex determination signals control ovo-B transcription in Drosophila melanogaster germ cells.

Nonautonomous inductive signals from the soma and autonomous signals due to a 2X karyotype determine the sex of Drosophila melanogaster germ cells. These two signals have partially overlapping influences on downstream sex determination genes. The upstream OVO-B transcription factor is required for the viability of 2X germ cells, regardless of sexual identity, and for female germline sexual identity. The influence of inductive and autonomous signals on ovo expression has been controversial. We show that ovo-B is strongly expressed in the 2X germ cells in either a male or a female soma. This indicates that a 2X karyotype controls ovo-B expression in the absence of inductive signals from the female soma. However, we also show that female inductive signals positively regulate ovo-B transcription in the 1X germ cells that do not require ovo-B function. Genetic analysis clearly indicates that inductive signals from the soma are not required for ovo-B function in 2X germ cells. Thus, while somatic inductive signals and chromosome karyotype have overlapping regulatory influences, a 2X karyotype is a critical germline autonomous determinant of ovo-B function in the germline.

Animals↗