Extraction of RNA and Analysis of Estrogen-Responsive Genes by RT-qPCR.

Reverse transcription-quantitative RT-PCR (RT-qPCR) is a powerful tool for assessing gene transcription levels. The technique is especially useful for measuring estrogen receptor transcript levels as well as gene expression changes in response to estrogen stimulation as it is quick, accurate, and robust and allows the measurement of gene expression in a variety of tissues and cells. This chapter describes the protocols used for RNA extraction and analysis as well as for RT-qPCR assay using hydrolysis (TaqMan-type) probes.

Protocol for DNA Microarrays on Glass Slides.

The DNA microarray is a powerful, flexible, nonbiased discovery technology. Microarrays can be used to assess processes from gene expression to long noncoding RNAs to specific pathologies, as well as many others. This chapter describes the protocol for DNA microarray analysis of differential gene expression using DNA sequences spotted on microscope slides.

Strain-specific differences in brain gene expression in a hydrocephalic mouse model with motile cilia dysfunction.

Congenital hydrocephalus results from cerebrospinal fluid accumulation in the ventricles of the brain and causes severe neurological damage, but the underlying causes are not well understood. It is associated with several syndromes, including primary ciliary dyskinesia (PCD), which is caused by dysfunction of motile cilia. We previously demonstrated that mouse models of PCD lacking ciliary proteins CFAP221, CFAP54 and SPEF2 all have hydrocephalus with a strain-dependent severity.

The Streptococcus pyogenes fibronectin/tenascin-binding protein PrtF.2 contributes to virulence in an influenza superinfection.

Influenza A virus (IAV) and Streptococcus pyogenes (the group A Streptococcus; GAS) are important contributors to viral-bacterial superinfections, which result from incompletely defined mechanisms. We identified changes in gene expression following IAV infection of A549 cells. Changes included an increase in transcripts encoding proteins with fibronectin-type III (FnIII) domains, such as fibronectin (Fn), tenascin N (TNN), and tenascin C (TNC).

Context-dependent regulation of pectoralis myostatin and lipid transporters by temperature and photoperiod in dark-eyed juncos.

A prominent example of seasonal phenotypic flexibility is the winter increase in thermogenic capacity (=summit metabolism, [Formula: see text]) in small birds, which is often accompanied by increases in pectoralis muscle mass and lipid catabolic capacity. Temperature or photoperiod may be drivers of the winter phenotype, but their relative impacts on muscle remodeling or lipid transport pathways are little known. We examined photoperiod and temperature effects on pectoralis muscle expression of myostatin, a muscle growth inhibitor, and its tolloid-like protein activators (TLL-1 and TLL-2), and sarcolemmal and intracellular lipid transporters in dark-eyed juncos .