Giant taxon-character matrices: the future of morphological systematics.

Simões et al. () argued that large matrices are linked to the construction of "problematic" characters, and that those characters negatively affect tree topology. In their re-evaluation of two squamate datasets, however, Simões et al.

High-throughput purification of hexahistidine-tagged proteins expressed in E. coli.

This chapter describes a method for efficient high-throughput purification of hexahistidine-tagged proteins that are expressed in Escherichia coli (E. coli) using immobilized metal affinity chromatography (IMAC) in a 96-well format. This approach is particularly suitable for proteomic applications that require modest amounts of highly purified proteins to be generated very efficiently.

Screening for the expression of soluble recombinant protein in Escherichia coli.

Protein expression and purification have traditionally been time-consuming, case-specific endeavors, and are considered to be the greatest bottlenecks in most proteomics pipelines. Escherichia coli (E. coli) is the most convenient and cost-effective host, although optimal conditions for the expression of different proteins vary widely.

High-throughput cloning for proteomics research.

Ligation-independent cloning (LIC) is a simple, rapid, and efficient method for high-throughput cloning. In this system, linear plasmid vector and insert DNA are treated to generate complementary single-stranded overhangs that anneal during a short incubation. The LIC system is adaptable for use with any vector following an alteration of the vector sequence.

Specificity of protein interactions mediated by BRCT domains of the XRCC1 DNA repair protein.

Protein interactions critical to DNA repair and cell cycle control systems are often coordinated by modules that belong to a superfamily of structurally conserved BRCT domains. Because the mechanisms of BRCT interactions and their significance are not well understood, we sought to define the affinity and specificity of those BRCT modules that orchestrate base excision repair and single-strand break repair. Common to these pathways is the essential XRCC1 DNA repair protein, which interacts with at least nine other proteins and DNA.

Fluffy, the major regulator of conidiation in Neurospora crassa, directly activates a developmentally regulated hydrophobin gene.

The fluffy (fl) gene of Neurospora crassa is required for asexual sporulation and encodes an 88 kDa polypeptide containing a typical fungal Zn2Cys6 DNA-binding motif. Identification of genes regulated by fl will provide insight into how fungi regulate growth during morphogenesis. As a step towards identifying the target genes on which FL may act, we sought to define target sequences to which the FL protein binds.

A saturation screen for cis-acting regulatory DNA in the Hox genes of Ciona intestinalis.

A screen for the systematic identification of cis-regulatory elements within large (>100 kb) genomic domains containing Hox genes was performed by using the basal chordate Ciona intestinalis. Randomly generated DNA fragments from bacterial artificial chromosomes containing two clusters of Hox genes were inserted into a vector upstream of a minimal promoter and lacZ reporter gene. A total of 222 resultant fusion genes were separately electroporated into fertilized eggs, and their regulatory activities were monitored in larvae.

A quality mix: using evidence and experience to evaluate new technologies.

This article describes BlueCross BlueShield of Tennessee's (BCBST) success in blending evidence with experience. This process has provided a winning solution for members, providers, and the health plan. Provider input has proved to be an invaluable tool in assessing both new and established technologies.

An improved method for the in vitro evolution of aptamers and applications in protein detection and purification.

One of the key components of proteomics initiatives is the production of high affinity ligands or probes that specifically recognize protein targets in assays that detect and capture proteins of interest. Particularly versatile probes with tremendous potential for use as affinity molecules are aptamers. Aptamers are short single-stranded DNA or RNA sequences that are selected in vitro based on affinity for a target molecule.

High-throughput proteomics: a flexible and efficient pipeline for protein production.

Many studies that aim to characterize the proteome require the production of pure protein in a high-throughput format. We have developed a system for high-throughput subcloning, protein expression and purification that is simple, fast, and inexpensive. We utilized ligation-independent cloning with a custom-designed vector and developed an expression screen to test multiple parameters for optimal protein production in E.

The draft genome of Ciona intestinalis: insights into chordate and vertebrate origins.

The first chordates appear in the fossil record at the time of the Cambrian explosion, nearly 550 million years ago. The modern ascidian tadpole represents a plausible approximation to these ancestral chordates. To illuminate the origins of chordate and vertebrates, we generated a draft of the protein-coding portion of the genome of the most studied ascidian, Ciona intestinalis.