Hybridization probe

In molecular biology, a hybridization probe is a fragment of DNA or RNA of variable length (usually 100-1000 bases long), which is used in DNA or RNA samples to detect the presence of nucleotide sequences (the DNA target) that are complementary to the sequence in the probe. The probe thereby hybridizes to single-stranded nucleic acid (DNA or RNA) whose base sequence allows probe-target base pairing due to complementarity between the probe and target. The labeled probe is first denatured (by heating or under alkaline conditions such as exposure to sodium hydroxide) into single stranded DNA (ssDNA) and then hybridized to the target ssDNA (Southern blotting) or RNA (northern blotting) immobilized on a membrane or in situ.

To detect hybridization of the probe to its target sequence, the probe is tagged (or labelled) with a molecular marker of either radioactive or (more recently) fluorescent molecules; commonly used markers are ³²P (a radioactive isotope of phosphorus incorporated into the phosphodiester bond in the probe DNA) or Digoxigenin, which is non-radioactive antibody-based marker. DNA sequences or RNA transcripts that have moderate to high sequence similarity to the probe are then detected by visualizing the hybridized probe via autoradiography or other imaging techniques. Normally, either X-ray pictures are taken of the filter, or the filter is placed under UV light. Detection of sequences with moderate or high similarity depends on how stringent the hybridization conditions were applied — high stringency, such as high hybridization temperature and low salt in hybridization buffers, permits only hybridization between nucleic acid sequences that are highly similar, whereas low stringency, such as lower temperature and high salt, allows hybridization when the sequences are less similar. Hybridization probes used in DNA microarrays refer to DNA covalently attached to an inert surface, such as coated glass slides or gene chips, and to which a mobile cDNA target is hybridized.

Depending on the method the probe may be synthesized using phosphoramidite method or generated and labeled by PCR amplification or cloning (older methods). In order to increase the in vivo stability of the probe RNA is not used, instead RNA analogues may be used, in particular morpholino. Molecular DNA- or RNA-based probes are now routinely used in screening gene libraries, detecting nucleotide sequences with blotting methods, and in other gene technologies like microarrays.

Examples of probes

Scorpion® probes

Molecular Beacon probes

TaqMan® probes

LNA® (Locked Nucleic Acid)probes

Cycling Probe Technology (CPT)

Use in forensic science

In forensic science, hybridization probes are used, for example, for detection of short tandem repeats (microsatellite) regions and in RFLP methods, all of which are widely used as part of DNA profiling analysis.

This biotechnology article is a stub. You can help Wikipedia by expanding it.v · Categories: Molecular biology Biotechnology stubs Wikimedia Foundation. 2010. Игры ⚽ Нужен реферат? Dharma transmission Punk Core Records Look at other dictionaries: Probe — may refer to:Media and entertainment* Probe (1972 TV pilot), a pilot for the sci fi series Search * Probe (TV series), a 1988 series * Probe (Philippine TV series) * Probe Records, a record label * Probe Entertainment, a British videogame… …   Wikipedia Hybridization, in situ — The use of a DNA or RNA probe to detect the complementary sequence. In situ hybridization is like all nucleic acid hybridization in being a technique in which single stranded nucleic acids (DNA or RNA) are permitted to interact so that complexes …   Medical dictionary Probe — (1) In surgery, a probe is a slender flexible rod with a blunt end used to explore, for example, an opening to see where it goes. (2) In molecular genetics, a probe is a labeled bit of DNA or RNA used to find its complementary sequence or locate… …   Medical dictionary hybridization — 1. The process of breeding a hybrid. 2. Crossing over between related but nonallelic genes. 3. The specific reassociation of complementary strands of polynucleic acids; e.g., the formation of a DNA RNA hybrid. 4. The process or act of forming a… …   Medical dictionary probe — A labelled DNA or RNA sequence used to detect the presence of a complementary sequence by hybridization with a nucleic acid sample …   Glossary of Biotechnology Nucleic acid hybridization — Hybridization is the process, discovered by Alexander Rich, of combining complementary, single stranded nucleic acids into a single molecule. Nucleotides will bind to their complement under normal conditions, so two perfectly complementary… …   Wikipedia Molecular Inversion Probe — (MIP)[1] belongs to the class of Capture by Circularization molecular techniques [1] for performing genomic partitioning, a process through which one captures and enriches specific regions of the genome[2]. Probes used in this technique are… …   Wikipedia Fluorescence in situ hybridization — A metaphase cell positive for the bcr/abl rearrangement (associated with chronic myelogenous leukemia) using FISH. The chromosomes can be seen in blue. The chromosome that is labeled with green and red spots (upper left) is the one where the… …   Wikipedia Fluorescent in situ hybridization — can be used to find out where the fluorescent probe bound to the chromosome. FISH is often used for finding specific features in DNA. These features can be used in genetic counseling, medicine, and species identification. Probes Probes are often… …   Wikipedia In situ hybridization — (ISH) is a type of hybridization that uses a labeled complementary DNA or RNA strand (i.e., probe) to localize a specific DNA or RNA sequence in a portion or section of tissue ( in situ ), or, if the tissue is small enough (e.g. plant seeds,… …   Wikipedia 18+ © Academic, 2000-2024 Contact us: Technical Support, Advertising Dictionaries export, created on PHP, Joomla, Drupal, WordPress, MODx. Mark and share Search through all dictionaries Translate… Search Internet Share the article and excerpts Direct link … Do a right-click on the link above and select “Copy Link”