PubMed Health⌕ Search

PubMed · 17018151

Design of a combinatorial DNA microarray for protein-DNA interaction studies.

Abstract

BACKGROUND: Discovery of precise specificity of transcription factors is an important step on the way to understanding the complex mechanisms of gene regulation in eukaryotes. Recently, double-stranded protein-binding microarrays were developed as a potentially scalable approach to tackle transcription factor binding site identification. RESULTS: Here we present an algorithmic approach to experimental design of a microarray that allows for testing full specificity of a transcription factor binding to all possible DNA binding sites of a given length, with optimally efficient use of the array. This design is universal, works for any factor that binds a sequence motif and is not species-specific. Furthermore, simulation results show that data produced with the designed arrays is easier to analyze and would result in more precise identification of binding sites. CONCLUSION: In this study, we present a design of a double stranded DNA microarray for protein-DNA interaction studies and show that our algorithm allows optimally efficient use of the arrays for this purpose. We believe such a design will prove useful for transcription factor binding site identification and other biological problems.

Explore related subjects

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Julian Mintseris, Michael B Eisen. 2006-10-03. Design of a combinatorial DNA microarray for protein-DNA interaction studies.. https://doi.org/10.1186/1471-2105-7-429

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related citations

A sensitive one-step real-time RT-PCR method for detecting Grapevine leafroll-associated virus 2 variants in grapevine.

Grapevine leafroll syndrome is caused by a complex of up to nine different Grapevine leafroll-associated viruses (GLRaV-1-9) with GLRaV-2 being reported as one of the most variable species of this group. Many methods, including indexing, serological and molecular procedures, have been developed for the detection of GLRaV-2. However, due to the low concentration of the virus in plants and the high variability of GLRaV-2, a method with improved sensitivity and with the capacity to detect of all known variants is required. Such improvement is essential for grapevine rootstocks, as these are suspected to harbour frequent GLRaV-2 infections difficult to detect, thus contributing to the spread of the leafroll disease. The development of new universal primers is described using a target sequence located in the 3' end of the virus genome. These primers were combined with a one-step SYBR Green real-time RT-PCR assay to achieve quantitative detection. All 43 GLRaV-2 isolates tested in this study were identified readily and reproducibly, regardless of their geographical origin or variety of grapevine. Using the procedure developed in this study, the sensitivity was increased 125 times compared to a conventional single-tube RT-PCR. This real-time method opens new perspectives for the sanitary selection of grapevine and in leafroll 2 disease monitoring.

3' Flanking Region↗

The presence of the internalin gene in natural atypically hemolytic Listeria innocua strains suggests descent from L. monocytogenes.

The atypical hemolytic Listeria innocua strains PRL/NW 15B95 and J1-023 were previously shown to contain gene clusters analogous to the pathogenicity island (LIPI-1) present in the related foodborne gram-positive facultative intracellular pathogen Listeria monocytogenes, which causes listeriosis. LIPI-1 includes the hemolysin gene, thus explaining the hemolytic activity of the atypical L. innocua strains. No other L. monocytogenes-specific virulence genes were found to be present. In order to investigate whether any other specific L. monocytogenes genes could be identified, a global approach using a Listeria biodiversity DNA array was applied. According to the hybridization results, the isolates were defined as L. innocua strains containing LIPI-1. Surprisingly, evidence for the presence of the L. monocytogenes-specific inlA gene, previously thought to be absent, was obtained. The inlA gene codes for the InlA protein which enables bacterial entry into some nonprofessional phagocytic cells. PCR and sequence analysis of this region revealed that the flanking genes of the inlA gene at the upstream, 5'-end region were similar to genes found in L. monocytogenes serotype 4b isolates, whereas the organization of the downstream, 3'-end region was similar to that typical of L. innocua. Sequencing of the inlA region identified a small stretch reminiscent of the inlB gene of L. monocytogenes. The presence of two clusters of L. monocytogenes-specific genes makes it unlikely that PRL/NW 15B95 and J1-023 are L. innocua strains altered by horizontal transfer. It is more likely that they are distinct relics of the evolution of L. innocua from an ancestral L. monocytogenes, as postulated by others.

3' Flanking Region↗

Development of the light-harvesting chlorophyll antenna in the green alga Chlamydomonas reinhardtii is regulated by the novel Tla1 gene.

The Chlamydomonas reinhardtii tla1 (truncated light-harvesting chlorophyll antenna size) mutant was generated upon DNA insertional mutagenesis and shown to specifically possess a smaller than wild type (WT) chlorophyll antenna size in both photosystems. Molecular and genetic analysis revealed that the exogenous plasmid DNA was inserted at the end of the 5' UTR and just prior to the ATG start codon of a hitherto unknown nuclear gene (termed Tla1), which encodes a protein of 213 amino acids. The Tla1 gene in the mutant is transcribed with a new 5' UTR sequence, derived from the 3' end of the transforming plasmid. This replacement of the native 5' UTR and promoter regions resulted in enhanced transcription of the tla1 gene in the mutant but inhibition in the translation of the respective tla1 mRNA. Transformation of the tla1 mutant with WT Tla1 genomic DNA successfully rescued the mutant. These results are evidence that polymorphism in the 5' UTR of the Tla1 transcripts resulted in the tla1 phenotype and that expression of the Tla1 gene is a prerequisite for the development/assembly of the Chl antenna in C. reinhardtii. A blast search with the Tla1 deduced amino acid sequence

3' Flanking Region↗