A. Daily Blog
Section I
The steps of gene cloning
Vectors in Molecular Cloning
Vectors are a type of DNA that acts as a carrier of a DNA segment that is to be cloned. In comparison to a chromosome, vectors could possibly carry just one gene while chromosomes can carry thousands of genes. The vector is a derivative of plasmids and viruses. Plasmids are small circular pieces of DNA that can be sed as vectors in genetic cloning. Viral vectors are a type of vector derived from viruses. Restriction enzymes then bind to specific DNA sequences and cleave the DNA at certain points on the vector and on the gene of interest. When a gene of interest and vector are cleaved by the same restriction endonuclease, then they can bind to each other resulting in a recombinant vector. Finally, the membranes of the target cells are made permeable to DNA allowing the recombinant vectors to enter. In summation, the vector is a glorified tupperware container.
Genomic vs cDNA
Genomic libraries are a type of DNA library in which the inserts are derived from chromosomal DNA.
Complementary DNA (cDNA) libraries are a type of DNA library in which the inserts are derived from cDNA. cDNA are DNA molecules made from mRNA. This means that the cDNA lacks the introns that cDNA has.
Think of the cDNA library as the SparkNotes version of the genomic library. While the genomic library is the original work and contains all of the genetic information, the cDNA library is a brief summary.
DNA Fragments and Gel Electrophoresis
Gel electrophoresis is the use of electrical charges to measure the amount of base pairs in a DNA fragment. The DNA is loaded on the negative side of the chamber and is attracted to the positive side of the chamber because DNA is naturally negatively charged.
Section II
Genomics vs Functional Genomics
Genomics are techniques that are used in the molecular analysis of the entire genome of a species. While genomics is aimed at studying the structure and function of many genes as a large integrated network, functional genomics is a genomic method aimed at studying the expression of the genome.
In Vitro DNA Synthesis Techniques
DNA Microarrays
DNa microarrays are a type of technology used to monitor the expression of thousands of genes simultaneously. Oligonucleotide probes are arranged on a small solid support surface. They represent the nucleotide sequences in known genes. The position of each probe is known. Microarrays can help identify the organisms and specific genes by hybridization of DNA from the organism to the oligonucleotide probes in the microarray.
Microarrays are really helpful in biological and biomedical research because the presence od antibiotic resistance, gene expression, and various other traits can be monitored with DNA microarrays.
Section III
Examples of Medically Important Eukaryotic Proteins Produced in Bacteria
Human Proteins in Sheep Cells? What?
A plasmid vector with the human protein is introduced to the sheep cell. It makes its way inside the sheep cell and integrates with the rest of the genome. Then, the gene is passed on to the offspring.
Applying Cloning Technology
Genetically Modified Organisms
These are organisms that carry genes that were introduced using molecular techniques such as gene cloning. They can either result in gene replacement or gene addition. Gene replacement is a phenomenon in which a cloned gene recombines with the normal gene on a chromosome and replaces it. Gene addition is the insertion of a cloned gene into the genome of an organism.
Transgenic Organism vs. Gene Knock-Out Manipulation
Transgenic organisms carry genes that were introduced using molecular techniques such as gene cloning.
Gene knock-out is when an organism's two copies of the functional gene have been replaced with he nonfunctional copies. Eventually, this can occur via gene replacement. While gene knock-out is a loss of function, transgenic organisms gain new capabilities.
Gene Cloning vs. Reproductive Cloning
Gene cloning is the process of making multiple copies of a gene of interest.
Reproductive cloning is the cloning of a multicellular organism, like a plant or an animal.
Bacteria respond to different environmental conditions that they may face, which is something that people knew. For instance, growth and cell division are monitored to ensure that the metabolism stays balanced and the cell lives. Alot of studies are now showing that the toxin-antitoxin system acts are metabolism regulators when bacteria are in a stressful environment. In addition, those studies show a potential correlation between the TA system and the antibiotic resistance of the bacteria and the stability of mobile genetic material. What is really up to debate now are the evolutionary origins of this and all of the correlations. They know that there could be a correlation but they don't know what that correlation is. Lastly, the TA system's physiology in bacteria had a big effect on cloning genes that are difficult to maintain, plasmid maintenance, and the production of recombinant proteins.
This is a diagram of gene cloning in bacteria. As you can see, a plasmid vector is used in this example. | |
This is a picture of a DNA microarray. Take note of the different fluorescent colors used. |