Answer:
Background
During the course of a bacterial infection, the rapid identification of the causative agent(s) is necessary for the determination of effective treatment options. We have developed a method based on a modified broad-range PCR and an oligonucleotide microarray for the simultaneous detection and identification of 12 bacterial pathogens at the species level. The broad-range PCR primer mixture was designed using conserved regions of the bacterial topoisomerase genes gyrB and parE. The primer design allowed the use of a novel DNA amplification method, which produced labeled, single-stranded DNA suitable for microarray hybridization. The probes on the microarray were designed from the alignments of species- or genus-specific variable regions of the gyrB and parE genes flanked by the primers. We included mecA-specific primers and probes in the same assay to indicate the presence of methicillin resistance in the bacterial species. The feasibility of this assay in routine diagnostic testing was evaluated using 146 blood culture positive and 40 blood culture negative samples.
Explanation:
Results
Comparison of our results with those of a conventional culture-based method revealed a sensitivity of 96% (initial sensitivity of 82%) and specificity of 98%. Furthermore, only one cross-reaction was observed upon investigating 102 culture isolates from 70 untargeted bacteria. The total assay time was only three hours, including the time required for the DNA extraction, PCR and microarray steps in sequence.
Transcription factors are necessary for an initiation of transcription at a regulated gene but not sufficient.
Transcription is the first step of gene expression in which DNA molecule is copied (transcribed) into RNA (mRNA) by RNA polymerase. The process of transcription is divided into three phases:
1. Initiation
• RNA polymerase with transcriptional factors bind to gene promoter Transcription factors can enhance the interaction between RNA polymerase and a DNA sequence- promoter, encouraging the expression of the gene. Such transcription factors are called activators. Otherwise, when the gene expression is inhibited, factors are called repressors and they bind to sequence –operator.
• RNA polymerase unwinds DNA double helix (transcription bubble is formed)
2. Elongation
• RNA polymerases adds nucleotides complementary to DNA
3. Termination
• RNA polymerase gets to stop codon (transcribes a sequence of DNA known as a terminator)
• Formed complementary RNA strand is released from DNA-RNA complex
