DNA Tech-Genomics-Proteomics Flashcards
Biotechnology
any technique applied to biological systems to manipulate processes for a specific purpose
DNA technologies
isolate, purify, analyze and manipulate DNA sequences
Genetic engineering
uses DNA technologies to alter genes for practical purposes
Cloning DNA
break open cells, let chromosomes come out, get the plasmid out of bacteria and cut open the plasmid using restriction enzymes (the same restriction enzyme to cut DNA and to open plasmid), use DNA ligase to attach the pieces of DNA with a plasmid
Bacterial plasmids that are engineered to contain:
- gene of interest
2. sorting genes
How can we isolate the bacteria that has our gene of interest?
Make agar plate and add ampocilin, if the bacteria doesn’t have the AMP gene it will die. Add X-gal substrate for the enzyme beta-galaticylase, if beta breaks down X-gal it turns blue (meaning it has the gene), even if it has the gene there can not be a gap in between
How to further isolate the gene of interest?
use a nucleic acid probe to identify gene of interest in set of clones. Break open the cells and denature them (make single stranded), add a single stranded DNA probe that only matches the gene of interest and add some form of dye. Observe which pot the gene is in
Genomic library
collection of clones containing every sequence in a genome
Complementary DNA (cDNA) library
DNA sequences made from expressed RNA
Make cDNA
go back into cell, build a complementary DNA strand using reverse transcriptase, add a complementary strand onto mRNA, every single mRNA has a polyAtail so that we have the primer and reverse transcriptase. Get rid of the RNA strand (using enzymes) and add DNA polymerase to do DNA replication
Polymerase chain reaction
denaturation, annealing, replication, taq polymerase. Denature DNA by increasing temp. add a DNA polymerase and nucleotides to start replicating DNA. Need to add taq polymerase to withstand. Repeat at least 30 times
Restriction fragments length polymorophisms
different fragments from the same part of genome
Southern blot analysis
uses electrophoresis, blot transfer, and labelled probes to identify RFLPs
DNA fingerprinting
observing short tandem repeats
-parts of nucleotide sequence that is repeated over and over
Transgenic organisms
modified to contain genes (trangene) from external source
Animal genetic engineering
use germ-line insert transgene, allow to grow and divide, inject into an embryo, place in a surrogate mouse. Some will have transgene, some wont
Stem Cell
unspecialized cell that can:
- reproduce indefinitely
- differentiate into various specialized cells
Embryonic stem cells
isolated from early embryos at the blastocyst stage
-can differentiate into any cell types (pluripotent)
Adult stem cells
replace nonreproducing specialized cells
Gene therapy
alteration of individual’s genes. May treat disorders with one defective gene
Dolly
take a cell from mammory gland, get an egg from another ewe and remove the nucleus and fuse the new nucleus into the egg and implant it into a third sheep as a surrogate
Genomics
is the characterization of whole genomes (structure, function, evolution)
Sequencing
determine the nucleotide sequence of a genome
Annotation
analyses specific regions of nucleotide regions
