Biology Genetic Engineering Gel Electrophoresis Juliana Machado Isabella Rojas Gabriel Felipe Galvis

Genetic Engineering Also called genetic modification. Is the human manipulation of an organisms genome (hereditary information of the organism) using modern DNA technology. It involves the introduction of DNA or synthetic genes into the organism.

History  Humans have altered the genomes of species for thousands of years through artificial methods. Genetic engineering considered the direct manipulation of DNA by humans has only existed in 1970s.  Paul Berg created the firsr recombinant ( DNA sequences that result from the use of laboratory methods). By combining the monkey virus SV40 with lambda phage (another virus consisting of a head)

 Herbert and Stanley Cohen created the first transgenic organism by inserting antibiotic resistance genes into the plasmid of an E. Coli bacterium.  Rudolf Jaenisch created a transgenic mouse by introducing DNA into its embryo, making it the first transgenic animal.  In 1976, Genentech, the first engineering company was founded by Herbert Boyer and Robert Swanson.

Applications Genetic engineering has applications in medicine, research, industry and agriculture. Medicine: Genetic engineering has been used to produce insulin, human growth hormones. Also it is used to create animal models which are non-human animals, used during the research and investigation of human diseases. Genetically modified mice are the most common genetically engineered animal model.

 Research: Genetic engineering is an important tool for natural scientists. Genes and other genetic information from a wide range of organisms and information are transformed into bacteria for storage and modification, creating genetically modified bacteria in the process. Also organisms are genetically engineered to discover the functions of certain genes. This could be the effect on the phenotype of the organism, where the gene is expressed or what other genes it interacts with.

 Industry: By engineering genes it is possible to create a biological factory that can produce proteins and enzymes. Bacteria and yeast have been used to produce medicines such as insulin, human growth and vaccines ( biological preparation that improves immunity to a particular disease).

 Agriculture: Genetic engineering is used in the creation of genetically food. Genetically modified organisms have had specific changes introduced into their DNA by genetic engineering techniques. These techniques are much more precise, an example is mutagenesis (mutation breeding) where an organism is exposed to radiation or chemicals to create a non- specific but stable change. Other techniques by which humans modify organisms include: selective breeding, plant breeding and animal breeding. Since genetically modified food has been introduced into supermarkets, there has been much controversy.

What is Gel electrophoresis? When DNA manipulation began, DNA fragments were separated by gravity. In the 1970s, the powerful tool of DNA electrophoresis was developed. Gel electrophoresis in chemistry: Is a method used to separate proteins by charge and size Gel electrophoresis in biochemistry: In molecular biology it is used to separate DNA and RNA fragments by length, to estimate the size of DNA and RNA.

Electricity effects on molecules  Nucleic acids molecules are separated by using a electric field to move the negatively charged molecules through an agarose matrix  We need to take into account that shorter molecules move faster and migrate longer than longer ones because shorter molecules migrate more easily through the pores of the gel. This process is called sieving. is a simple and convenient technique of separating particles of different size  In the case of proteins they are separated by charge in agarose becuase the pores of the gel are too large to sieve proteins Also electrophoresis can be used in the separation of nanoparticles.

Agarose  Agarose is a polysacharide obtained from agar that is used for a variety of science applications, specially in gel electrophoresis.  Agarose it is used in the matrix, which plays a vital role in this separation.

Agarose used as a gel to be incluided in the matrix.

How does gel electrophoresis work? - After DNA is extracted from cells or obtained using PCR (polymerase chain reaction), they are cut into many fragments of different lenghts with restriction enzymes (enzyme that cuts DNA).

-DNA fragments are loaded into the wells of an agarose gel and placed in an electrophoresis buffer filled tank. This gel which is similar to gelatin is formed for the DNA fragments to be placed in here. -This gel is placed in the matrix and contains electrolytes, so that an electric current conducted. -Positive and negative electrodes are connected to the opposite ends of the gel- (electric field is applied). -The negatively charged DNA fragments are drawn towards the positive terminal.

 The agar gel is highly porous  Shortest DNA fragments are able to move quickly through the pores.  Larger DNA fragments move very slowly because it is more difficult for them to pass through the pores.

Applications of Gel electrophoresis  Estimation of the size of DNA molecules. Example: restriction enzyme (enzyme that cuts DNA) also called restriction mapping of cloned DNA.  Analysis of PCR(polymerase chain reaction) products is a scientific technique in molecular biology to amplify or increase a single or few copies of a piece of DNA across several orders, generating thousands or millions of copies of a particular DNA sequence. 8 PCR tubes, each containing a reaction mixture

What is DNA profiling?  DNA profiling also called DNA fingerprinting, DNA testing, is a technique use by forensic scientists to distinguish between individuals of the same species using samples of their DNA.  Although the majority of our DNA is the same, there are short pieces called microsatellites. These repeat many times in a persons DNA. This is the pattern of repeats which is different in every individual.  Knowing these microsatellites, DNA sequences are the basis for the DNA profiling.

Stages of DNA profiling  STEP 1: Cells are broken down to release DNA. If a small amount of DNA is available, the amount can be increased or amplified by using polymerase chain reaction (PCR).  STEP 2: The DNA is cut into fragments using restrictions enzymes as I explained before. Each restriction enzyme cuts DNA at a specific base sequence.  The sections of DNA which are cut are called restriction fragments.

 STEP 3: In DNA profiling the gel electrophoresis is involved Fragments are separated on the basis by using a process called gel electrophoresis.  DNA fragments are inserted in the wells and a electric current is developed. As I explained before in gel electrophoresis.  DNA is negatively charged so it is attracted to the positive part of the gel.  DNA is separated on the basis of size.  A radioactive is added which combines with the DNA fragments to produce a image, that contains the copies of DNA bands.  STEP 4: The pattern of the fragment is analyzed.

USES OF DNA PROFILING  DNA profiling is used to solve medical problems.  Biological uses for DNA profiling: - Blood - Hair - Saliva - Body tissue cells

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