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Transcription, Genetic: The biosynthesis of RNA carried out on a template of DNA. The biosynthesis of DNA from an RNA template is called Reverse transcription.

Transcription

JoVE 10794

Transcription is the process of synthesizing RNA from a DNA sequence by RNA polymerase. It is the first step in producing a protein from a gene sequence. Additionally, many other proteins and regulatory sequences are involved in the proper synthesis of messenger RNA (mRNA). Regulation of transcription is responsible for the differentiation of all the different types of cells and often for the proper cellular response to environmental signals. In eukaryotes, the DNA is first transcribed into a primary RNA, or pre-mRNA, that can be further processed into a mature mRNA to serve as a template for the synthesis of proteins. In prokaryotes such as bacteria, however, translation of RNA into polypeptides can begin while the transcription is still ongoing, as RNA can be quickly degraded. Transcription can also produce different kinds RNA molecules that do not code for protein, such as microRNAs, transfer RNA (tRNA), and ribosomal RNA (rRNA)—all of which contribute to protein synthesis. With few exceptions, all of the cells in the human body have the same genetic information in them, from neurons in the brain to muscle cells in the heart. So how do cells assume such diverse forms and functions? To a large extent, the answer lies in the regulation of transcription during development of the organism. Specifically, transcriptional regulation plays a central ro

 Core: Biology

What is Gene Expression?

JoVE 10797

Gene expression is the process in which DNA directs the synthesis of functional products, such as proteins. Cells can regulate gene expression at various stages. It allows organisms to generate different cell types and enables cells to adapt to internal and external factors.

A gene is a stretch of DNA that serves as the blueprint for functional RNAs and proteins. Since DNA is made up of nucleotides and proteins consist of amino acids, a mediator is required to convert the information that is encoded in DNA into proteins. This mediator is the messenger RNA (mRNA). mRNA copies the blueprint from DNA by a process called transcription. In eukaryotes, transcription takes place in the nucleus by complementary base-pairing with the DNA template. The mRNA is then processed and transported into the cytoplasm where it serves as a template for protein synthesis during translation. In prokaryotes, which lack a nucleus, the processes of transcription and translation occur at the same location and almost simultaneously since the newly-formed mRNA is susceptible to rapid degradation. Every cell of an organism contains the same DNA, and consequently the same set of genes. However, not all genes in a cell are “turned on” or use to synthesize proteins. A gene is said to be “expressed” when the protein it encodes is produced by the cell. Gen

 Core: Biology

Translation

JoVE 10795

Translation is the process of synthesizing proteins from the genetic information carried by messenger RNA (mRNA). Following transcription, it constitutes the final step in the expression of genes. This process is carried out by ribosomes, complexes of protein and specialized RNA molecules. Ribosomes, transfer RNA (tRNA) and other proteins are involved in the production of the chain of amino acids—the polypeptide. Proteins are called the “building blocks” of life because they make up the vast majority of all organisms—from muscle fibers to hairs on your head to components of your immune system—and the blueprint for each and every one of those proteins is encoded by the genes found in the DNA of every cell. The central dogma in biology dictates that genetic information is converted into functional proteins by the processes of transcription and translation. Eukaryotes have a membrane-bound nucleus where mRNA is transcribed from DNA. After transcription, mRNA is shuttled out of the nucleus to be translated into a chain of amino acids—a polypeptide—and eventually, a functional protein. This can take place in the cytoplasm or in the rough endoplasmic reticulum, where the polypeptides are further modified. By contrast, prokaryotes lack a nuclear compartment, so translation in prokaryotes takes place in the cytoplasm,

 Core: Biology

Operons

JoVE 10984

Prokaryotes can control gene expression through operons—DNA sequences consisting of regulatory elements and clustered, functionally related protein-coding genes. Operons use a single promoter sequence to initiate transcription of a gene cluster (i.e., a group of structural genes) into a single mRNA molecule. The terminator sequence ends transcription. An operator sequence, located between the promoter and structural genes, prohibits the operon’s transcriptional activity if bound by a repressor protein. Altogether, the promoter, operator, structural genes, and terminator form the core of an operon. Operons are usually either inducible or repressible. Inducible operons, such as the bacterial lac operon, are normally “off” but will turn “on” in the presence of a small molecule called an inducer (e.g., allolactose). When glucose is absent, but lactose is present, allolactose binds and inactivates the lac operon repressor—allowing the operon to generate enzymes responsible for lactose metabolism. Repressible operons, such as the bacterial trp operon, are usually “on” but will turn “off” in the presence of a small molecule called a corepressor (e.g., tryptophan). When tryptophan—an essential amino acid—is abundant, tryptophan binds and activates the

 Core: Biology

The Central Dogma

JoVE 10798

The central dogma of biology states that information encoded in the DNA is transferred to messenger RNA (mRNA), which then directs the synthesis of protein. The set of instructions that enable the mRNA nucleotide sequence to be decoded into amino acids is called the genetic code. The universal nature of this genetic code has spurred advances in scientific research, agriculture, and medicine. In the early 1900s, scientists discovered that DNA stores all the information needed for cellular functions and that proteins perform most of these functions. However, the mechanisms of converting genetic information into functional proteins remained unknown for many years. Initially, it was believed that a single gene is directly converted into its encoded protein. Two crucial discoveries in eukaryotic cells challenged this theory: First, protein production does not take place in the nucleus. Second, DNA is not present outside the nucleus. These findings sparked the search for an intermediary molecule that connects DNA with protein production. This intermediary molecule, found in both the nucleus and the cytoplasm, and associated with protein production, is RNA. During transcription, RNA is synthesized in the nucleus, using DNA as a template. The newly-synthesized RNA is similar in sequence to the DNA strand, except thymidine in DNA is replaced by uracil i

 Core: Biology

Organization of Genes

JoVE 10786

The genomes of eukaryotes can be structured in several functional categories. A strand of DNA is comprised of genes and intergenic regions. Genes themselves consist of protein-coding exons and non-coding introns. Introns are excised once the sequence is transcribed to mRNA, leaving only exons to code for proteins.

In eukaryotic genomes, genes are separated by large stretches of DNA that do not code for proteins. However, these intergenic regions carry important elements that regulate gene activity, for instance, the promoter where transcription starts, and enhancers and silencers that fine-tune gene expression. Sometimes these binding sites can be located far away from the associated gene. As researchers investigated the process of gene transcription in eukaryotes, they realized that the final mRNA that codes for a protein is shorter than the DNA it is derived from. This difference in length is due to a process called splicing. Once pre-mRNA has been transcribed from DNA in the nucleus, splicing immediately removes introns and joins exons together. The result is protein-coding mRNA that moves to the cytoplasm and is translated into protein. One of the largest human genes, DMD, is over two million base pairs long. This gene encodes the muscle protein dystrophin. Mutations in DMD cause muscular dystrophy, a disorder characteri

 Core: Biology

Epigenetic Regulation

JoVE 10803

Epigenetic mechanisms play an essential role in healthy development. Conversely, precisely regulated epigenetic mechanisms are disrupted in diseases like cancer.

In most mammals, females have two X chromosomes (XX) while males have an X and a Y chromosome (XY). The X chromosome contains significantly more genes than the Y chromosome. Therefore, to prevent an excess of X chromosome-linked gene expression in females, one of the two X chromosomes is randomly silenced during early development. This process, called X-chromosome inactivation, is regulated by DNA methylation. Scientists have found greater DNA methylation at gene promoter sites on the inactive X chromosome than its active counterpart. DNA methylation prevents the transcription machinery from attaching to the promoter region, thus inhibiting gene transcription. Abnormal DNA methylation plays an important role in cancer. The promoter region of most genes contains stretches of cytosine and guanine nucleotides linked by a phosphate group. These regions are called CpG islands. In healthy cells, CpG islands are not methylated. However, in cancer cells, CpG islands in the promoter regions of tumor suppressor genes or cell cycle regulators are excessively methylated. Methylation turns off the expression of these genes, allowing cancer cells to divide rapidly and uncontrollably.

 Core: Biology

Bacterial Transformation- Concept

JoVE 10573

Background

In early 20th century, pneumonia was accountable for a large portion of infectious disease deaths1. In order to develop an effective vaccine against pneumonia, Frederick Griffith set out to study two different strains of the Streptococcus pneumoniae: a non-virulent strain with a rough appearance (R-strain) and a virulent strain with a smooth appearance…

 Lab Bio

RNA Analysis of Environmental Samples Using RT-PCR

JoVE 10104

Source: Laboratories of Dr. Ian Pepper and Dr. Charles Gerba - Arizona University
Demonstrating Author: Bradley Schmitz


Reverse transcription-polymerase chain reaction (RT-PCR) involves the same process as conventional PCR — cycling temperature to amplify nucleic acids. However, while conventional PCR only amplifies…

 Environmental Microbiology

The Nucleus

JoVE 10691

The nucleus is a membrane-bound organelle that contains a eukaryotic organism’s genetic instructions in the form of chromosomal DNA. This is distinct from the DNA in mitochondria or chloroplasts that carry out functions specific to those organelles. While some cells—such as red blood cells—do not have a nucleus, and others—such as skeletal muscle cells—have multiple nuclei, most eukaryotic cells have a single nucleus. The DNA in the nucleus is wrapped around proteins such as histones, creating a DNA-protein complex called chromatin. When cells are not dividing—that is, when they are in the interphase part of their cell cycle—the chromatin is organized diffusely. This allows easy access to the DNA during the transcription process when messenger RNA (mRNA) is synthesized based on the DNA code. When a eukaryotic cell is about to divide, the chromatin condenses tightly into distinct, linear chromosomes. Humans have 46 chromosomes in total. Chromatin is particularly concentrated in a region of the nucleus called the nucleolus. The nucleolus is important for the production of ribosomes, which translate mRNA into protein. In the nucleolus, ribosomal RNA is synthesized and combined with proteins to create ribosomal subunits, which later form functioning ribosomes in the cytoplasm of the cell. The interior of t

 Core: Biology
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