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Expression Cloning
Expression cloning is a technique in DNA cloning that uses expression vectors to generate a library of clones, with each clone expressing one protein. This ''expression library'' is then screened for the property of interest and clones of interest are recovered for further analysis. An example would be using an expression library to isolate genes that could confer antibiotic resistance. Expression vectors Expression vectors are a specialized type of cloning vector in which the transcriptional and translational signals needed for the regulation of the gene of interest are included in the cloning vector. The transcriptional and translational signals may be synthetically created to make the expression of the gene of interest easier to regulate. Purpose Usually the ultimate aim of expression cloning is to produce large quantities of specific proteins. To this end, a bacterial expression clone may include a ribosome binding site ( Shine-Dalgarno sequence) to enhance translation of the ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
DNA Cloning
Molecular cloning is a set of experimental methods in molecular biology that are used to assemble recombinant DNA molecules and to direct their replication within host organisms. The use of the word ''cloning'' refers to the fact that the method involves the replication of one molecule to produce a population of cells with identical DNA molecules. Molecular cloning generally uses DNA sequences from two different organisms: the species that is the source of the DNA to be cloned, and the species that will serve as the living host for replication of the recombinant DNA. Molecular cloning methods are central to many contemporary areas of modern biology and medicine. In a conventional molecular cloning experiment, the DNA to be cloned is obtained from an organism of interest, then treated with enzymes in the test tube to generate smaller DNA fragments. Subsequently, these fragments are then combined with vector DNA to generate recombinant DNA molecules. The recombinant DNA is then ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Transcription (genetics)
Transcription is the process of copying a segment of DNA into RNA for the purpose of gene expression. Some segments of DNA are transcribed into RNA molecules that can encode proteins, called messenger RNA (mRNA). Other segments of DNA are transcribed into RNA molecules called non-coding RNAs (ncRNAs). Both DNA and RNA are nucleic acids, which use base pairs of nucleotides as a complementary language. During transcription, a DNA sequence is read by an RNA polymerase, which produces a complementary, antiparallel RNA strand called a primary transcript. In virology, the term transcription is used when referring to mRNA synthesis from a viral RNA molecule. The genome of many RNA viruses is composed of negative-sense RNA which acts as a template for positive sense viral messenger RNA - a necessary step in the synthesis of viral proteins needed for viral replication. This process is catalyzed by a viral RNA dependent RNA polymerase. Background A DNA transcription unit encoding ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
PBR322
pBR322 is a plasmid and was one of the first widely used ''E. coli'' cloning vectors. Created in 1977 in the laboratory of Herbert Boyer at the University of California, San Francisco, it was named after Francisco Bolivar Zapata, the postdoctoral researcher and Raymond L. Rodriguez. The p stands for "plasmid," and BR for "Bolivar" and "Rodriguez." pBR322 is 4361 base pairs in length and has two antibiotic resistance genes – the gene '' bla'' encoding the ampicillin resistance (AmpR) protein, and the gene ''tetA'' encoding the tetracycline resistance (TetR) protein. It contains the origin of replication of pMB1, and the '' rop'' gene, which encodes a restrictor of plasmid copy number. The plasmid has unique restriction sites for more than forty restriction enzymes. Eleven of these forty sites lie within the TetR gene. There are two sites for restriction enzymes HindIII and ClaI within the promoter of the TetR gene. There are six key restriction sites inside the AmpR gene ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
λ Phage
Lambda phage (coliphage λ, scientific name ''Lambdavirus lambda'') is a bacterial virus, or bacteriophage, that infects the bacterial species ''Escherichia coli'' (''E. coli''). It was discovered by Esther Lederberg in 1950. The wild type of this virus has a Temperate (virology), temperate life cycle that allows it to either reside within the genome of its host through lysogeny or enter into a lytic phase, during which it kills and lyses the cell to produce offspring. Lambda strains, mutated at specific sites, are unable to lysogenize cells; instead, they grow and enter the lytic cycle after superinfecting an already lysogenized cell. The phage particle consists of a head (also known as a capsid), a tail, and tail fibers (see image of virus below). The head contains the phage's double-strand linear DNA genome. During infections, the phage particle recognizes and binds to its host, ''E. coli'', causing DNA in the head of the phage to be ejected through the tail into the cytopla ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Translation (biology)
In biology, translation is the process in living Cell (biology), cells in which proteins are produced using RNA molecules as templates. The generated protein is a sequence of amino acids. This sequence is determined by the sequence of nucleotides in the RNA. The nucleotides are considered three at a time. Each such triple results in the addition of one specific amino acid to the protein being generated. The matching from nucleotide triple to amino acid is called the genetic code. The translation is performed by a large complex of functional RNA and proteins called ribosomes. The entire process is called gene expression. In translation, messenger RNA (mRNA) is decoded in a ribosome, outside the nucleus, to produce a specific amino acid chain, or polypeptide. The polypeptide later protein folding, folds into an Activation energy, active protein and performs its functions in the cell. The polypeptide can also start folding during protein synthesis. The ribosome facilitates decoding ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Gene Expression
Gene expression is the process (including its Regulation of gene expression, regulation) by which information from a gene is used in the synthesis of a functional gene product that enables it to produce end products, proteins or non-coding RNA, and ultimately affect a phenotype. These products are often proteins, but in non-protein-coding genes such as Transfer RNA, transfer RNA (tRNA) and Small nuclear RNA, small nuclear RNA (snRNA), the product is a functional List of RNAs, non-coding RNA. The process of gene expression is used by all known life—eukaryotes (including multicellular organisms), prokaryotes (bacteria and archaea), and viruses—to generate the macromolecule, macromolecular machinery for life. In genetics, gene expression is the most fundamental level at which the genotype gives rise to the phenotype, ''i.e.'' observable trait. The genetic information stored in DNA represents the genotype, whereas the phenotype results from the "interpretation" of that informati ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Genetics
Genetics is the study of genes, genetic variation, and heredity in organisms.Hartl D, Jones E (2005) It is an important branch in biology because heredity is vital to organisms' evolution. Gregor Mendel, a Moravian Augustinians, Augustinian friar working in the 19th century in Brno, was the first to study genetics scientifically. Mendel studied "trait inheritance", patterns in the way traits are handed down from parents to offspring over time. He observed that organisms (pea plants) inherit traits by way of discrete "units of inheritance". This term, still used today, is a somewhat ambiguous definition of what is referred to as a gene. Phenotypic trait, Trait inheritance and Molecular genetics, molecular inheritance mechanisms of genes are still primary principles of genetics in the 21st century, but modern genetics has expanded to study the function and behavior of genes. Gene structure and function, variation, and distribution are studied within the context of the Cell (bi ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Posttranslational Modification
In molecular biology, post-translational modification (PTM) is the covalent process of changing proteins following protein biosynthesis. PTMs may involve enzymes or occur spontaneously. Proteins are created by ribosomes, which translate mRNA into polypeptide chains, which may then change to form the mature protein product. PTMs are important components in cell signalling, as for example when prohormones are converted to hormones. Post-translational modifications can occur on the amino acid side chains or at the protein's C- or N- termini. They can expand the chemical set of the 22 amino acids by changing an existing functional group or adding a new one such as phosphate. Phosphorylation is highly effective for controlling the enzyme activity and is the most common change after translation. Many eukaryotic and prokaryotic proteins also have carbohydrate molecules attached to them in a process called glycosylation, which can promote protein folding and improve stability a ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Eukaryotes
The eukaryotes ( ) constitute the domain of Eukaryota or Eukarya, organisms whose cells have a membrane-bound nucleus. All animals, plants, fungi, seaweeds, and many unicellular organisms are eukaryotes. They constitute a major group of life forms alongside the two groups of prokaryotes: the Bacteria and the Archaea. Eukaryotes represent a small minority of the number of organisms, but given their generally much larger size, their collective global biomass is much larger than that of prokaryotes. The eukaryotes emerged within the archaeal kingdom Promethearchaeati and its sole phylum Promethearchaeota. This implies that there are only two domains of life, Bacteria and Archaea, with eukaryotes incorporated among the Archaea. Eukaryotes first emerged during the Paleoproterozoic, likely as flagellated cells. The leading evolutionary theory is they were created by symbiogenesis between an anaerobic Promethearchaeati archaean and an aerobic proteobacterium, which form ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Expression Vector
An expression vector, otherwise known as an expression construct, is usually a plasmid or virus designed for gene expression in cells. The vector (molecular biology), vector is used to introduce a specific gene into a target cell, and can commandeer the cell's mechanism for protein synthesis to produce the protein Genetic code, encoded by the gene. Expression vectors are the basic tools in biotechnology for the protein production, production of proteins. The Vector (molecular biology), vector is engineered to contain regulatory sequences that act as Enhancer (genetics), enhancer and Promoter (biology), promoter regions and lead to efficient transcription of the gene carried on the expression vector. The goal of a well-designed expression vector is the efficient production of protein, and this may be achieved by the production of significant amount of stable messenger RNA, which can then be Translation (biology), translated into protein. The expression of a protein may be tightly ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Binding Site
In biochemistry and molecular biology, a binding site is a region on a macromolecule such as a protein that binds to another molecule with specificity. The binding partner of the macromolecule is often referred to as a ligand. Ligands may include other proteins (resulting in a protein–protein interaction), enzyme substrates, second messengers, hormones, or allosteric modulators. The binding event is often, but not always, accompanied by a conformational change that alters the protein's function. Binding to protein binding sites is most often reversible (transient and non-covalent), but can also be covalent reversible or irreversible. Function Binding of a ligand to a binding site on protein often triggers a change in conformation in the protein and results in altered cellular function. Hence binding site on protein are critical parts of signal transduction pathways. Types of ligands include neurotransmitters, toxins, neuropeptides, and steroid hormones. Binding site ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
