Molecular biology
Molecular biology is a branch of biology that seeks to understand the molecular structures and chemical processes underlying biological activity within and between cells. It centers largely on nucleic acids such as DNA and RNA, and on proteins. In 1945, the English physicist William Astbury reached for a new term to describe an emerging way of seeing life. He called it molecular biology, an approach focused on the physical and chemical structures of biological molecules. Astbury wanted to explain the observations of classical biology, which studied living processes at larger scales and higher levels of organization, by looking at the molecules themselves. Cells had been observed in organisms as early as the 18th century. Yet a detailed understanding of the mechanisms governing their behavior did not arrive until the 20th century. What did scientists have to discover before life could be read at the level of its molecules? How did a dye, a kitchen blender, and a heavier kind of nitrogen each rewrite what we knew about inheritance? And how do researchers today copy a single strand of DNA into more than a billion?
In 1953, Francis Crick, James Watson, Rosalind Franklin, and their colleagues at the Medical Research Council Unit, Cavendish Laboratory, were the first to describe the double helix model for the chemical structure of deoxyribonucleic acid. This is often considered a landmark event for the young field. It provided a physico-chemical basis for understanding the previously nebulous idea of nucleic acids as the primary substance of biological inheritance. Watson and Crick built their model on earlier research by Franklin, whose X-ray crystallography work was conveyed to them by Maurice Wilkins and Max Perutz. Their proposed structure let them conjecture about how DNA might replicate. Watson and Crick described not only the shape but the implications of that shape. The recognition followed years later. 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. From this single structure flowed the discovery of DNA in other microorganisms, plants, and animals.
In 1866, Gregor Mendel first described the laws of inheritance he observed in his studies of mating crosses in pea plants. His work gave the field its name, Mendelian genetics. One of his laws, the law of segregation, states that diploid individuals with two alleles for a particular gene pass one of these alleles to their offspring. The chemistry came into view in 1869, when the Swiss biochemist Friedrich Miescher proposed a structure he called nuclein, which we now know to be DNA. Miescher found this substance by studying the components of pus-filled bandages, noting the unique properties of the phosphorus-containing substances. In 1919, Phoebus Levene proposed the polynucleotide model of DNA, drawing on his biochemical experiments on yeast. In 1950, Erwin Chargaff expanded on Levene's work and elucidated critical properties of nucleic acids. The sequence of nucleic acids varies across species. And the total concentration of purines, adenine and guanine, always equals the total concentration of pyrimidines, cytosine and thymine. This balance is now known as Chargaff's rule, and it would point the way toward the helix itself.
In 1928, Frederick Griffith encountered a virulence property in pneumococcus bacteria that was killing lab rats. According to Mendel, prevalent at that time, gene transfer could occur only from parent to daughter cells. Griffith advanced another theory, that gene transfer occurring in members of the same generation, now called horizontal gene transfer, was taking place. Griffith worked with two strains: one virulent and smooth, one avirulent and rough. The smooth strain had a glistering appearance from a specific polysaccharide capsule, a polymer of glucose and glucuronic acid. That capsule hid the bacteria from the host's immune system, letting it kill the host. The rough strain lacked the capsule and was readily destroyed. In 1944, Oswald Avery, Colin MacLeod, and Maclyn McCarty reported that DNA is the substance that causes bacterial transformation. This came in an era when it was widely believed that proteins carried genetic information. Their work culminated research from the 1930s at the Rockefeller Institute for Medical Research to purify the transforming principle first described in Griffith's experiment. In their paper in the February 1944 issue of the Journal of Experimental Medicine, they suggested that DNA, rather than protein, may be the hereditary material of bacteria. The decisive confirmation came from the Hershey-Chase experiment. Alfred Hershey and Martha Chase used E. coli and bacteriophage, and a kitchen blender as a major piece of apparatus. They tagged the phage's protein coat with radioactive sulfur and its DNA with radioactive phosphorus in two separate test tubes. After mixing, blending to separate phage from cells, and centrifuging, the E. coli cells showed radioactive phosphorus. The transformed material was DNA, not the protein coat, and that finding gave rise to the theory of transduction.
In 1958, Matthew Meselson and Franklin Stahl ran an experiment that supported Watson and Crick's hypothesis that DNA replication was semiconservative. In semiconservative replication, each of the two new double-stranded helices consists of one strand from the original helix and one newly synthesized strand. The experiment has been called the most beautiful experiment in biology. Meselson and Stahl decided the best way to trace the parent DNA would be to tag it by changing one of its atoms. Since nitrogen is present in all of the DNA bases, they grew parent DNA containing a heavier isotope of nitrogen than occurs naturally. That altered mass let them determine how much parent DNA remained after successive cycles of replication. The same period sharpened the reading of the code itself. In 1961, it was demonstrated that three sequential bases of a gene's DNA specify each successive amino acid of a protein. The genetic code is a triplet code, where each triplet, called a codon, specifies a particular amino acid. The codons do not overlap, and each sequence is read from a fixed starting point. During 1962 to 1964, conditional lethal mutants of a bacterial virus drove fundamental advances in understanding the proteins of DNA replication, repair, recombination, and assembly.
Polymerase chain reaction, or PCR, is an extremely versatile technique for copying DNA. Under perfect conditions it could amplify one DNA molecule into 1.07 billion molecules in less than two hours. PCR is used to study gene expression, detect pathogenic microorganisms, find genetic mutations, and introduce mutations through site-directed mutagenesis. Its variations include reverse transcription PCR for amplifying RNA and quantitative PCR for measuring DNA or RNA molecules. Molecular cloning, a recombinant DNA technology first developed in the 1960s, isolates a DNA sequence and transfers it into a plasmid vector. The plasmid usually has three distinctive features: an origin of replication, a multiple cloning site, and a selective marker, usually antibiotic resistance. DNA can enter bacterial cells by transformation, conjugation, or transduction, while introducing DNA into eukaryotic cells by physical or chemical means is called transfection. Gel electrophoresis separates molecules by size using an agarose or polyacrylamide gel. Because the DNA backbone carries negatively charged phosphate groups, DNA migrates toward the positive end of the current. Proteins can be separated by size using an SDS-PAGE gel. The Bradford assay, developed in 1975 by Marion M. Bradford, quantitates protein using a dye called Coomassie Brilliant Blue G-250. When the dye binds protein in an acidic solution, its background wavelength shifts from 465 nm to 595 nm, turning from reddish-brown to bright blue. Proteins bind the dye in about 2 minutes, and readings are recommended within 5 to 20 minutes. Unlike the older Lowry procedure and biuret assay, the Bradford assay resists interference from ethanol, sodium chloride, and magnesium chloride, though strong alkaline agents such as sodium dodecyl sulfate can disturb it.
Edwin Southern gave his name to the Southern blot, a method for probing for a specific DNA sequence within a sample. DNA samples are separated by gel electrophoresis, transferred to a membrane by blotting via capillary action, and exposed to a labeled DNA probe with a complementary base sequence. The names northern, western, and eastern blotting all grew from a molecular biology joke that played on Southern blotting. Patricia Thomas developed the RNA blot that became known as the northern blot, though she did not actually use the term. The northern blot studies specific RNA molecules and is commonly used to learn at what time and under what conditions certain genes are expressed in living tissues. A western blot detects specific proteins from a mixture, first separating them by size through SDS-PAGE, then transferring them to a membrane such as polyvinylidene fluoride or nitrocellulose, which is probed with antibodies. The eastern blotting technique detects post-translational modification of proteins. A DNA microarray is a collection of spots on a solid support such as a microscope slide, each spot holding single-stranded DNA oligonucleotide fragments about 100 micrometre in diameter. Arrays can compare the gene expression of two different tissues, such as a healthy and a cancerous one, and can carry anywhere from 100 spots to more than 10,000. Allele-specific oligonucleotide detects single base mutations using short probes 20 to 25 nucleotides in length, without the need for PCR or gel electrophoresis. Older methods have given way to these: before DNA gel electrophoresis, the size of DNA molecules was measured by rate sedimentation in sucrose gradients, and before that, by viscometry.
Molecular biology is multi-disciplinary, drawing on genetics, biochemistry, physics, mathematics, and more recently computer science through bioinformatics. It sits at the intersection of biochemistry and genetics, both of which seek the molecular mechanisms that underlie vital cellular functions. Biochemistry studies the chemical substances and vital processes in living organisms, focusing on biomolecules such as proteins, lipids, carbohydrates, and nucleic acids. Genetics studies how genetic differences affect organisms and attempts to predict how mutations and genetic interactions shape the expression of a phenotype. Molecular genetics, the study of gene structure and function, has been among the most prominent sub-fields since the early 2000s. Much of molecular biology is quantitative, and a significant amount of work now uses computer science techniques such as bioinformatics and computational biology. In the early 2020s, the field entered a golden age defined by both vertical and horizontal technical development. Vertically, novel technologies allow real-time monitoring of biological processes at the atomic level. Horizontally, sequencing data is becoming more affordable across many scientific fields, driving development in developing nations. CRISPR-Cas9 gene editing experiments can now be conceived and implemented by individuals for under $10,000 in novel organisms, opening a path toward industrial and medical applications that earlier generations could only imagine.
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Common questions
What is molecular biology?
Molecular biology is a branch of biology that seeks to understand the molecular structures and chemical processes underlying biological activity within and between cells. It centers largely on nucleic acids such as DNA and RNA, and on proteins, examining how they orchestrate processes like replication, transcription, and translation.
Who first used the term molecular biology?
The English physicist William Astbury first used the term molecular biology in 1945. He described it as an approach focused on uncovering the physical and chemical structures of biological molecules and how their interactions explain the observations of classical biology.
Who described the double helix structure of DNA and when?
In 1953, Francis Crick, James Watson, Rosalind Franklin, and their colleagues at the Medical Research Council Unit, Cavendish Laboratory, were the first to describe the double helix model for the chemical structure of DNA. Watson and Crick were awarded the Nobel Prize in Physiology or Medicine in 1962, along with Maurice Wilkins.
What did the Hershey-Chase experiment prove about DNA?
The Hershey-Chase experiment confirmed that DNA is the genetic material that causes infection. Alfred Hershey and Martha Chase tagged a bacteriophage's protein coat with radioactive sulfur and its DNA with radioactive phosphorus, and found that E. coli cells showed radioactive phosphorus, proving the transformed material was DNA, not protein.
How does PCR work in molecular biology?
Polymerase chain reaction, or PCR, is a technique for copying DNA that allows a specific sequence to be copied or modified. Under perfect conditions it could amplify one DNA molecule into 1.07 billion molecules in less than two hours, and it is used to study gene expression, detect pathogens, and introduce mutations.
What is the Bradford assay used for in molecular biology?
The Bradford assay enables fast, accurate quantitation of protein molecules using a dye called Coomassie Brilliant Blue G-250, which shifts from reddish-brown to bright blue upon binding protein. It was developed in 1975 by Marion M. Bradford and resists interference from molecules such as ethanol, sodium chloride, and magnesium chloride.