Module 1.2: DNA Recombinant Technology
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Recombinant DNA are DNA molecules formed by laboratory methods of ____ (i.e., molecular cloning) in order to piece together genetic material from ____ sources
genetic recombination; multiple
True or False. Recombinant DNA can be found naturally in the genome of organisms in nature.
False.
Recombinant DNA technology comprises altering genetic material ____ an organism to obtain _____ characteristics in living organisms or as their products.
outside; enhanced and desired
Recombinant DNA technology involves the insertion of ____ from a variety of sources, having a desirable gene sequence via the appropriate_____.
DNA fragments; vector
Match the proponent/s for the discovery in the history of rRNA technology. • Was awarded the 1986 Nobel Prize for medicine in his discoveries on growth factors. • Developed the first recombinant DNA utilizing bacterial DNA and plasmids. • Conceived and approach to create rDNAs in vitro • Created the first chimeric DNA in vitro • Isolated the new cloning vector, pSC101 and created bacterial intra- and interspecies rDNAs Choices: Berg and colleagues Berg and Loban Herbert Boyer Stanley Cohen Jackson and colleagues
• Stanley Cohen • Herbert Boyer and Stanley Cohen • Berg and Loban • Jackson and colleagues • Stanley Cohen and colleagues
What was the first plasmid cloning vector?
lambda DVGal 120 (an operon encoding galactose metabolism genes concatenated with simian vacoulating virus 40)
This hormone discovered by Stanley Cohen is involved in the proliferation and differentiation of cells in vivo.
Nerve growth factors and epidermal growth factors
Memorize/familiarize
• An approach to creating rDNA in vitro is conceived • The first bacterial cloning vector was isolated • First chimeric DNA is created • Second cloning vector is isolated • First intentional creation of rDNA molecules • Ribosomal genes propagated in E. coli cells • Asilomar conference looked at safety and containment issues of extrachromosomal DNAs and its risks of cancer • First gene transfer with humans • Gene therapy products are regulated by FDA • First approved gene transfer study on a chile with SCID • Gelsinger’s death • FDA elevates administrative unit that evaluates cellular, tissue, and gene therapy products.
Genetic engineering refers to the process of using ____ to modify an organism’s ____ to achieve ____.
recombinant DNA technology; DNA; desirable traits
Addition of ____ in the form of ____ that are generated by ____ is the most common method of genetic engineering
foreign DNA; recombinant DNA vectors; molecular cloning
Other terms that are synonymous with genetic engineering
Recombinant DNA technology Gene manipulation Gene modification
____ is the production of an exact copy—specifically, an exact genetic copy—of a gene, cell, or organism
DNA Cloning
____ allows for the creation of multiple copies of genes, expression of genes, and study of specific genes
Molecular cloning
_____ is a method used to make a clone or an identical copy of an entire multicellular organism.
Reproductive cloning
____ is a combination of DNA fragments generated by ____ that does not exist in nature.
Recombinant DNA; molecular cloning
Two types of enzymes in creating recombinant DNA.
restriction endonuclease; ligase
Pipeline for recombinant DNA technology
• Isolate the DNA sequence gene of interest • Insert DNA of interest into vector plasmid • Put recombinant plasmid into E. coli through the process of transformation • Clone cells • Choose colonies and inoculate a large culture to expand the plasmid carrying the gene of interest • Isolate and clean the DNA from the expanded culture • Transfer the plasmid DNA to desired cells and test the biological properties of the gene of interest
During DNA isolation for the rDNA experiments, DNA is coexisting with other macromolecules within the cell membrane; hence, it must be separated and purified using enzymes such as
lysozymes, cellulase, chitinase, ribonuclease, and proteases
During DNA isolation, DNA eventually precipitates out as fine threads as a result of the presence of ____.
ethanol
During the cutting of DNA for an rDNA experiment, ____ helps to identify the location wherein a designated gene is introduced into a vector genome. The said reaction is known as ____. ____ is a technology that displays the restriction enzyme digestion’s progress
Restriction enzymes; restriction enzyme digestion; ‘Agarose Gel Electrophoresis’
In the insertion to host phase, rDNA is added to the recipient host cell, and the entire process is called ____.
transformation (for bacteria) transfection (for eukaryotes) transduction (when a viral vector is used)
foreign DNA. If so, they are given treatments to make them ‘capable’ of accepting new DNA. Give an example of this treatment.
Calcium ion treatments. Ca-DNA complex, endocytosis, proteolytic degradation of the endosome containing Ca-DNA complex
In the recombinant cell isolation phase, only transformed host cells are filtered and isolated. The ___ of the plasmid vector is used to distinguish recombinant cells from non-recombinant cells.
marker gene
Sequence the basic steps in recombinant DNA technology: ____ Recombinant Isolation ____ DNA isolation ____ Amplifying DNA ____ Insertion of rRNA into a host ____ Cutting of DNA/Restriction enzyme digestion ____ Joining DNA
6, 1, 3, 5, 2, 4