Tuesday, October 4, 2011

Types of RNA

Transcription results in the production of RNA molecules, often generically referred to as transcripts. Traditionally, the transcripts observed within a cell were broadly classified as ribosomal RNA (rRNA), transfer RNA (tRNA), heterogeneous nuclear RNA (hnRNA), or messenger RNA (mRNA), as well as a collection of small RNAs of previously unknown function. Now, however, one must include the extremely diverse population of miRNAs and other RNAs which are of immense interest in the study of the regulation of gene expression . Each category of RNA, which is synthesized by a different type of RNA polymerase, performs a different function in the cell. These highly diverse populations of RNA are not represented in equal amounts in the cell and the relative amount of each is directly related to the physiology of the cell.
rRNA is the most abundant RNA component in the cell. In prokaryotic cells the major rRNA species are the 23S rRNA, 16S rRNA, and 5S rRNA. The eukaryotic counterparts are identified as the 28S rRNA, 18S rRNA, and 5S rRNA, as well as a fourth ribosomal transcript, the 5.8S rRNA. These molecules form the scaffolding of ribosomes, which become translationally competent when decorated with myriad ribosomal proteins. At present there are 55 known prokaryotic ribosomal proteins and 82 known eukaryotic ribosomal proteins. Not all ribosomes are functional at any given time, and the existence of a pool of transiently inactive ribosomes is itself a regulator of gene expression. The super abundance of rRNA is often exploited as both an RNA mass loading control as well as internal molecular weight markers for electrophoresis.
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Bioelectronic DNA detection involves forming an electronic circuit mediated by nucleic acid hybridization and it serves as the basis for a DNA detection system called eSensor™ [1-4]. This system uses low-density DNA chips containing electrodes coated with DNA capture probes. Target DNA present in the sample hybridizes specifically both to capture probes and ferrocene labeled signal probes in solution thereby generating an electric current. Currente Sensor DNA chips contain as many as 36 electrodes for simultaneous detection of multiple pathogens from a single sample.

Many pathogens cause both acute and chronic disease at relatively low copy number and may be difficult or impossible to propagate in culture. Thus, most pathogen detection systems rely on nucleic acid amplification by using polymerase chain reaction (PCR). One highly effective amplification strategy targets conserved sequences among the family of organisms of interest. Such broad-range PCR strategies have been used to identify and characterize several known and previously uncharacterized bacteria [5,6] and viruses [7,8]. In order to maximize the utility of these effective pathogen nucleic acid amplification systems, amplification needs to be coupled with rapid, sensitive, and specific detection. Bioelectronic DNA detection by use of the eSensor chip might fulfill this need.

Human papillomaviruses (HPV) serve as an ideal model system for determining the efficiency and feasibility of eSensor DNA detection technology since there are at least 30 distinct genital HPV types that can be effectively amplified with broad-range consensus PCR primers. We designed two eSensor chips, each containing 14 probes specific for the conserved L1 region of the HPV genome. We evaluated clinical cervical cytology samples known to contain one or more HPV types. The eSensor DNA detection platform successfully detected the correct HPV type in most of these clinical samples, demonstrating that the system provides a rapid, sensitive, specific, and economical approach for multiple-pathogen detection and identification from a single sample.Background We used human papillomaviruses (HPV) as a model system to evaluate the utility of a nucleic acid, hybridization-based bioelectronic DNA detection platform (eSensor) in identifying multiple pathogens.