Molecular biology
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Molecular biology /məˈlɛkjʊlər/ is the branch of biology that seeks to understand the molecular basis of biological activity in and between cells, including biomolecular synthesis, modification, mechanisms, and interactions.[1][2][3] The study of chemical and physical structure of biological macromolecules is known as molecular biology.[4]
Molecular biology was first described as an approach focused on the underpinnings of biological phenomena - uncovering the structures of biological molecules as well as their interactions, and how these interactions explain observations of classical biology.[5]
In 1945 the term molecular biology was used by physicist William Astbury. In 1953 Francis Crick, James Watson, Rosalind Franklin, and colleagues, working at Medical Research Council unit, Cavendish laboratory, Cambridge (now the MRC Laboratory of Molecular Biology), made a double helix model of DNA which changed the entire research scenario. They proposed the DNA structure based on previous research done by Rosalind Franklin and Maurice Wilkins. This research then lead to finding DNA material in other microorganisms, plants and animals.[4]
Molecular biology is not simply the study of biological molecules and their interactions; rather, it is also a collection of techniques developed since the field's genesis which have enabled scientists to learn about molecular processes.[6] In this way it has both complemented and improved biochemistry and genetics as methods (of understanding nature) that began before its advent. One notable technique which has revolutionized the field is the polymerase chain reaction (PCR), which was developed in 1983.[6] PCR is a reaction which amplifies small quantities of DNA, and it is used in many applications across scientific disciplines.[7][8]
The central dogma of molecular biology describes the process in which DNA is transcribed into RNA, which is then translated into protein.[2][9]
Molecular biology also plays a critical role in the understanding of structures, functions, and internal controls within individual cells, all of which can be used to efficiently target new drugs, diagnose disease, and better understand cell physiology.[10] Some clinical research and medical therapies arising from molecular biology are covered under gene therapy whereas the use of molecular biology or molecular cell biology in medicine is now referred to as molecular medicine.
History of molecular biology
Molecular biology sits at the intersection of biochemistry and genetics; as these scientific disciplines emerged and evolved in the 20th century, it became clear that they both sought to determine the molecular mechanisms which underlie vital cellular functions.[11] Advances in molecular biology have been closely related to the development of new technologies and their optimization.[12] Molecular biology has been elucidated by the work of many scientists, and thus the history of the field depends on an understanding of these scientists and their experiments.
The field of genetics arose as an attempt to understand the molecular mechanisms of genetic inheritance and the structure of a gene. Gregor Mendel pioneered this work in 1866, when he first wrote the laws of genetic inheritance based on his studies of mating crosses in pea plants.[13] One such law of genetic inheritance is the law of segregation, which states that diploid individuals with two alleles for a particular gene will pass one of these alleles to their offspring.[14] Because of his critical work, the study of genetic inheritance is commonly referred to as Mendelian genetics.[15]
A major milestone in molecular biology was the discovery of the structure of DNA. This work began in 1869 by Friedrich Miescher, a Swiss biochemist who first proposed a structure called nuclein, which we now know to be (deoxyribonucleic acid), or DNA.[16] He discovered this unique substance by studying the components of pus-filled bandages, and noting the unique properties of the "phosphorus-containing substances".[17] Another notable contributor to the DNA model was Phoebus Levene, who proposed the "polynucleotide model" of DNA in 1919 as a result of his biochemical experiments on yeast.[18] In 1950, Erwin Chargaff expanded on the work of Levene and elucidated a few critical properties of nucleic acids: first, the sequence of nucleic acids varies across species.[19] Second, the total concentration of purines (adenine and guanine) is always equal to the total concentration of pyrimidines (cysteine and thymine).[16] This is now known as Chargaff's rule. In 1953, James Watson and Francis Crick published the double helical structure of DNA,[20] using the X-ray crystallography work done by Rosalind Franklin and Maurice Wilkins. Watson and Crick described the structure of DNA and conjectured about the implications of this unique structure for possible mechanisms of DNA replication.[20]
Watson and Crick were awarded the Nobel Prize in Physiology or Medicine in 1962, along with Wilkins, for proposing a model of the structure of DNA.[4]
In 1961, it was demonstrated that when a gene encodes a protein, three sequential bases of a gene's DNA specify each successive amino acid of the protein.[21] Thus the genetic code is a triplet code, where each triplet (called a codon) specifies a particular amino acid. Furthermore, it was shown that the codons do not overlap with each other in the DNA sequence encoding a protein, and that each sequence is read from a fixed starting point.
During 1962–1964, through the use of conditional lethal mutants of a bacterial virus,
The F.Griffith experiment
In 1928, Fredrick Griffith, encountered a virulence property in pneumococcus bacteria, which was killing lab rats. According to Mendel, prevalent at that time, gene transfer could occur only from parent to daughter cells only. Griffith advanced another theory, stating that gene transfer occurring in member of same generation is known as horizontal gene transfer (HGT). This phenomenon is now referred to as genetic transformation.
Griffith addressed the Streptococcus pneumoniae bacteria, which had two different strains, one virulent and smooth and one avirulent and rough. The smooth strain had glistering appearance owing to the presence of a type of specific polysaccharide – a polymer of glucose and glucuronic acid capsule. Due to this polysaccharide layer of bacteria, a host's immune system cannot recognize the bacteria and it kills the host. The other, avirulent, rough strain lacks this polysaccharide capsule and has a dull, rough appearance.
Presence or absence of capsule in the strain, is known to be genetically determined. Smooth and rough strains occur in several different type such as S-I, S-II, S-III, etc. and R-I, R-II, R-III, etc. respectively. All this subtypes of S and R bacteria differ with each other in antigen type they produce.[4]
Hershey and Chase experiment
Confirmation that DNA is the genetic material which is cause of infection came from Hershey and Chase experiment. They used E.coli and bacteriophage for the experiment. This experiment is also known as blender experiment, as kitchen blender was used as a major piece of apparatus. Alfred Hershey and Martha Chase demonstrated that the DNA injected by a phage particle into a bacterium contains all information required to synthesize progeny phage particles. They used radioactivity to tag the bacteriophage's protein coat with radioactive sulphur and DNA with radioactive phosphorus, into two different test tubes respectively. After mixing bacteriophage and E.coli into the test tube, the incubation period starts in which phage transforms the genetic material in the E.coli cells. Then the mixture is blended or agitated, which separates the phage from E.coli cells. The whole mixture is centrifuged and the pellet which contains E.coli cells was checked and the supernatant was discarded. The E.coli cells showed radioactive phosphorus, which indicated that the transformed material was DNA not the protein coat.
The transformed DNA gets attached to the DNA of E.coli and radioactivity is only seen onto the bacteriophage's DNA. This mutated DNA can be passed to the next generation and the theory of Transduction came into existence. Transduction is a process in which the bacterial DNA carry the fragment of bacteriophages and pass it on the next generation. This is also a type of horizontal gene transfer.[4]
Modern molecular biology
In the early 2020s, molecular biology entered a golden age defined by both vertical and horizontal technical development. Vertically, novel technologies are allowing for real-time monitoring of biological processes at the atomic level.[23] Molecular biologists today have access to increasingly affordable sequencing data at increasingly higher depths, facilitating the development of novel genetic manipulation methods in new non-model organisms. Likewise, synthetic molecular biologists will drive the industrial production of small and macro molecules through the introduction of exogenous metabolic pathways in various prokaryotic and eukaryotic cell lines.[24]
Horizontally, sequencing data is becoming more affordable and used in many different scientific fields. This will drive the development of industries in developing nations and increase accessibility to individual researchers. Likewise, CRISPR-Cas9 gene editing experiments can now be conceived and implemented by individuals for under $10,000 in novel organisms, which will drive the development of industrial and medical applications [25]
Relationship to other biological sciences
and molecular biologyThe following list describes a viewpoint on the interdisciplinary relationships between molecular biology and other related fields.[26]
- Molecular biology is the study of the molecular underpinnings of the biological phenomena, focusing on molecular synthesis, modification, mechanisms and interactions.
- Biochemistry is the study of the chemical substances and vital processes occurring in living organisms. Biochemists focus heavily on the role, function, and structure of biomolecules such as proteins, lipids, carbohydrates and nucleic acids.[27]
- Genetics is the study of how genetic differences affect organisms. Genetics attempts to predict how mutations, individual genes and genetic interactions can affect the expression of a phenotype[28]
While researchers practice techniques specific to molecular biology, it is common to combine these with methods from genetics and biochemistry. Much of molecular biology is quantitative, and recently a significant amount of work has been done using computer science techniques such as bioinformatics and computational biology. Molecular genetics, the study of gene structure and function, has been among the most prominent sub-fields of molecular biology since the early 2000s. Other branches of biology are informed by molecular biology, by either directly studying the interactions of molecules in their own right such as in cell biology and developmental biology, or indirectly, where molecular techniques are used to infer historical attributes of populations or species, as in fields in evolutionary biology such as population genetics and phylogenetics. There is also a long tradition of studying biomolecules "from the ground up", or molecularly, in biophysics.[29]
Techniques of molecular biology
Molecular cloning
Molecular cloning is used to isolate and then transfer a DNA sequence of interest into a plasmid vector.[30] This recombinant DNA technology was first developed in the 1960s.[31] In this technique, a DNA sequence coding for a protein of interest is cloned using polymerase chain reaction (PCR), and/or restriction enzymes, into a plasmid (expression vector). The plasmid vector usually has at least 3 distinctive features: an origin of replication, a multiple cloning site (MCS), and a selective marker (usually antibiotic resistance). Additionally, upstream of the MCS are the promoter regions and the transcription start site, which regulate the expression of cloned gene.
This plasmid can be inserted into either bacterial or animal cells. Introducing DNA into bacterial cells can be done by transformation via uptake of naked DNA, conjugation via cell-cell contact or by transduction via viral vector. Introducing DNA into eukaryotic cells, such as animal cells, by physical or chemical means is called transfection. Several different transfection techniques are available, such as calcium phosphate transfection, electroporation, microinjection and liposome transfection. The plasmid may be integrated into the genome, resulting in a stable transfection, or may remain independent of the genome and expressed temporarily, called a transient transfection.[32][33]
DNA coding for a protein of interest is now inside a cell, and the protein can now be expressed. A variety of systems, such as inducible promoters and specific cell-signaling factors, are available to help express the protein of interest at high levels. Large quantities of a protein can then be extracted from the bacterial or eukaryotic cell. The protein can be tested for enzymatic activity under a variety of situations, the protein may be crystallized so its tertiary structure can be studied, or, in the pharmaceutical industry, the activity of new drugs against the protein can be studied.[34]
Polymerase chain reaction
in a gel tray.
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