Molecular biology is a branch of biology that seeks to understand the molecular basis of biological activity in and between cells, including biomolecular synthesis, modification, mechanisms, and interactions.
Oligonucleotides are short DNA or RNA molecules, oligomers, that have a wide range of applications in genetic testing, research, and forensics. Commonly made in the laboratory by solid-phase chemical synthesis, these small fragments of nucleic acids can be manufactured as single-stranded molecules with any user-specified sequence, and so are vital for artificial gene synthesis, polymerase chain reaction (PCR), DNA sequencing, molecular cloning and as molecular probes. In nature, oligonucleotides are usually found as small RNA molecules that function in the regulation of gene expression, or are degradation intermediates derived from the breakdown of larger nucleic acid molecules.
In molecular biology and biotechnology, a fluorescent tag, also known as a fluorescent label or fluorescent probe, is a molecule that is attached chemically to aid in the detection of a biomolecule such as a protein, antibody, or amino acid. Generally, fluorescent tagging, or labeling, uses a reactive derivative of a fluorescent molecule known as a fluorophore. The fluorophore selectively binds to a specific region or functional group on the target molecule and can be attached chemically or biologically. Various labeling techniques such as enzymatic labeling, protein labeling, and genetic labeling are widely utilized. Ethidium bromide, fluorescein and green fluorescent protein are common tags. The most commonly labelled molecules are antibodies, proteins, amino acids and peptides which are then used as specific probes for detection of a particular target.
A locked nucleic acid (LNA), also known as bridged nucleic acid (BNA), and often referred to as inaccessible RNA, is a modified RNA nucleotide in which the ribose moiety is modified with an extra bridge connecting the 2' oxygen and 4' carbon. The bridge "locks" the ribose in the 3'-endo (North) conformation, which is often found in the A-form duplexes. This structure provides for increased stability against enzymatic degradation. LNA also offers improved specificity and affinity in base-pairing as a monomer or a constituent of an oligonucleotide. LNA nucleotides can be mixed with DNA or RNA residues in a oligonucleotide.
Keratin, type I cytoskeletal 19 (Keratin-19)) also known as cytokeratin-19 (CK-19) is a 40 kDa protein that in humans is encoded by the KRT19 gene. Keratin-19 is a type I keratin.
In molecular biology, a hybridization probe (HP) is a fragment of DNA or RNA, usually 15–10000 nucleotides long, which can be radioactively or fluorescently labeled. HPs can be used to detect the presence of nucleotide sequences in analyzed RNA or DNA that are complementary to the sequence in the probe. The labeled probe is first denatured into single stranded DNA (ssDNA) and then hybridized to the target ssDNA or RNA immobilized on a membrane or in situ.
Cycling probe technology (CPT) is a molecular biological technique for detecting specific DNA sequences. CPT operates under isothermal conditions. In some applications, CPT offers an alternative to PCR. However, unlike PCR, CPT does not generate multiple copies of the target DNA itself, and the amplification of the signal is linear, in contrast to the exponential amplification of the target DNA in PCR. CPT uses a sequence specific chimeric probe which hybridizes to a complementary target DNA sequence and becomes a substrate for RNase H. Cleavage occurs at the RNA internucleotide linkages and results in dissociation of the probe from the target, thereby making it available for the next probe molecule. Integrated electrokinetic systems have been developed for use in CPT.
A real-time polymerase chain reaction is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR). It monitors the amplification of a targeted DNA molecule during the PCR, not at its end, as in conventional PCR. Real-time PCR can be used quantitatively and semi-quantitatively.
SYBR Green I (SG) is an asymmetrical cyanine dye used as a nucleic acid stain in molecular biology. The SYBR family of dyes is produced by Molecular Probes Inc., now owned by Thermo Fisher Scientific. SYBR Green I binds to DNA. The resulting DNA-dye-complex best absorbs 497 nanometer blue light and emits green light. The stain preferentially binds to double-stranded DNA, but will stain single-stranded (ss) DNA with lower performance. SYBR Green can also stain RNA with a lower performance than ssDNA.
Digital polymerase chain reaction is a biotechnological refinement of conventional polymerase chain reaction methods that can be used to directly quantify and clonally amplify nucleic acids strands including DNA, cDNA, or RNA. The key difference between dPCR and qPCR lies in the method of measuring nucleic acids amounts, with the former being a more precise method than PCR, though also more prone to error in the hands of inexperienced users. PCR carries out one reaction per single sample. dPCR also carries out a single reaction within a sample, however the sample is separated into a large number of partitions and the reaction is carried out in each partition individually. This separation allows a more reliable collection and sensitive measurement of nucleic acid amounts. The method has been demonstrated as useful for studying variations in gene sequences — such as copy number variants and point mutations.
The RNA integrity number (RIN) is an algorithm for assigning integrity values to RNA measurements.
A nucleic acid test (NAT) is a technique used to detect a particular nucleic acid sequence and thus usually to detect and identify a particular species or subspecies of organism, often a virus or bacterium that acts as a pathogen in blood, tissue, urine, etc. NATs differ from other tests in that they detect genetic materials rather than antigens or antibodies. Detection of genetic materials allows an early diagnosis of a disease because the detection of antigens and/or antibodies requires time for them to start appearing in the bloodstream. Since the amount of a certain genetic material is usually very small, many NATs include a step that amplifies the genetic material—that is, makes many copies of it. Such NATs are called nucleic acid amplification tests (NAATs). There are several ways of amplification, including polymerase chain reaction (PCR), strand displacement assay (SDA), transcription mediated assay (TMA), and loop-mediated isothermal amplification (LAMP).
The history of the polymerase chain reaction (PCR) has variously been described as a classic "Eureka!" moment, or as an example of cooperative teamwork between disparate researchers. Following is a list of events before, during, and after its development:
OLIGO Primer Analysis Software is a software for DNA primer design. The first paper describing this software was published in 1989. The program is a real time PCR primer and probe search and analysis tool. It additionally performs siRNA and molecular beacon searches, open reading frame analysis, and restriction enzyme analysis. It was created and maintained by Wojciech Rychlik and Piotr Rychlik.
MAGIChips, also known as "microarrays of gel-immobilized compounds on a chip" or "three-dimensional DNA microarrays", are devices for molecular hybridization produced by immobilizing oligonucleotides, DNA, enzymes, antibodies, and other compounds on a photopolymerized micromatrix of polyacrylamide gel pads of 100x100x20 μm or smaller size. This technology is used for analysis of nucleic acid hybridization, specific binding of DNA, and low-molecular weight compounds with proteins, and protein-protein interactions.
The FluoProbes series of fluorescent dyes were developed by Interchim to improve performances of standard fluorophores. They are designed for labeling biomolecules, cells, tissues or beads in advanced fluorescent detection techniques.
SYBR Safe is a cyanine dye used as a nucleic acid stain in molecular biology. SYBR Safe is one of a number of SYBR dyes made by the Life Technologies Corporation. SYBR Safe binds to DNA. The resulting DNA-dye-complex absorbs blue light and emits green light.
Tom Brown FRSC FRSE is a British chemist, biotechnologist, and entrepreneur. He is the Professor of Nucleic acid chemistry at the Department of Chemistry and Department of Oncology at the University of Oxford. Currently, he is serving as the President of the Chemical Biology Interface Division of the Royal Society of Chemistry. He is best known for his contribution in the field of DNA Repair, DNA Click chemistry, and in the application of Molecular genetics in forensics and diagnostics.
Pernilla Wittung-Stafshede is a Swedish biophysical chemist, born in 1968, who is a professor of chemical biology at Chalmers University of Technology in Gothenburg. In 2019 she was named by International Union of Pure and Applied Chemistry as a Distinguished Woman in Chemistry.
The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines are a set of protocols for conducting and reporting quantitative real-time PCR experiments and data, as devised by Bustin et al. in 2009. They were devised after a paper was published in 2002 that claimed to detect measles virus in children with autism through the use of RT-qPCR, but the results proved to be completely unreproducible by other scientists. The authors themselves also did not try to reproduce them and the raw data was found to have a large amount of errors and basic mistakes in analysis. This incident prompted Stephen Bustin to create the MIQE guidelines to provide a baseline level of quality for qPCR data published in scientific literature.
