Biotechnology (4th LE) Flashcards
Double stranded
Strands are complementary to each other
DNA
Base pairing of one strand of a nucleic acid to another nucleic acid
Nucleic acid hybridization
Laboratory techniques used to study and manipulate DNA
Used to analyze gene expression
DNA Technology
The manipulation of organisms or their components to make useful products
Biotechnology
Direct manipulation or genes for practical purposes
Cor of gene cloning
Genetic Engineering (Recombinant DNA Technology)
First automated procedure in sequencing
Sanger Method
Another name for sanger method
Dideoxyribonucleotuide (or dideoxy) chain terminating sequence
Developed the sanger method
Frederick Sanger
High throughput
Real-time result
Based on sequencing by synthesis
Next Generation DNA Sequencing
Used to synthesize a stretch of DNA from a single-stranded template, and the order in which nucleotides are added revels the sequence
DNA polymerase
Third generation
A single molecule of DNA or RNA can be sequenced without the need for PCR amplification or chemcial labeling of the sample
Nanopore Sequencing
Nanopore Sequencing principle
Each type or base interupts the electrical current for a slightly different length of time
A process of producing multiple copies of a gene dor the analysis and manipulation of DNA
Gene cloning (or DNA cloning)
The first to be performed during DNA manipulation
DNA Extraction
Used to rapidly amplify a specific target gene segment in a DNA
Polymerase Chain Reaction (PCR)
Introduced PCR
Kary Mullis in 1985
An equipment that enables the PCR reaction
Thermal cycler (PCR machine)
Steps in PCR
Heat denaturation at 94C
Cooling
Synthesis of new DNA
REPEAT
Usual number of cycles in PCR
30 cycles which would yield up to 10^9 fold increase
Two limitations of PCR
Sequences should be know otherwise primers cannot be made
Limit of DNA sequence that can be copied
Three techniques in studying PCR products
Gel electrophoresis
Cloning
Sequencing
Uses a gel to separate out a mixture or nucleic acid fragments by length/size
Gel Electrophoresis
Staining that makes DNA fragments clearly visible under UV
Ethidium Bromide (EtBr) staining
Small, circular DNA capable of independent replication
Plasmid
Used as a vector in recombinant DNA technology
Plasmid
Steps in gene cloning
Insertion of DNA Vector transports gene into host Multiplication within host Division of host cell A clone is produced
Recognize and cut DNA only at a particular sequence of nucleotides
Restriction Enzymes
Examples of restriction enzyme
EcoRI
Xbal
Xhol
Used in analyzing the result of restriction endonuclease cleavage
Gel electrophoresis
Seal the base-paired fragments (after restriction enzyme digestion) producing recombinant DNA molecules
DNA ligase
Uptake of DNA from the medium by bacterial cell
Use of heat shock
Transformation
DNA uptake through the use of electric shock
Electroporation
Equivalent to transformation, the only difference being that phage DNA rather than a plasmid is involved
Transfection
Introducing DNA into the cells that involve bombardment with high velocity microprojectiles coated with DNA
Biolistics
Another name for biolistics
Gene Gun
Cells without cell walls
Protoplast
Process that may be spontaneous or it may be induced
Protoplast Fusion
Method of introducing new DNA into a cell by injecting it directly into the nucleus
Microinjection
Recombinant DNA molecule can be inserted into a cell by:
Transformation Electroporation Transfection Biolistics Protoplast Fusion Microinjection
Methods to verify your gene of interest was successfully cloned
Blue-white screening
Restriction enzyme digestion
Colony PCR
DNA sequencing
Use bacterial lactose metabolism as an indicator
Blue-white screening
Color if the DNA insert is present
White
Color if the DNA insert is not present
Blue
Checks the presence and direction of your insert
Restriction enzyme digestion
The most rapid screening method in determining presence of the DNA insert
Colony PCR
The most accurate way
Verifies presence and exact sequence of inserted DNA
DNA Sequencing
The complete set of genes present in a cell or organism
Genome
Total mRNA molecules expressed from the genes of an organism in a given environmental condition
Transcriptome
Total proteins present in an organism under a given environmental condition
Proteome
A discipline in genetics that applies recombinant DNA, DNA sequencing methods, and bioinformatics to sequence, assemble, and analyze the function and structure of genomes
Genomics
Used to determine the regulatory potential of a DNA sequence that is unknown
Reporter Gene Assay
Used to detect specific RNA molecules present within an RNA mixture
Used to determine the RNA expression of certain genes
Northern Blotting
Used to detect specific protein molecules within a protein mixture
Can help in determining protein size and expression
Western Blotting
Used to visualize copy number aberrations such as the deletion, translocation or amplification of chromosomes
Fluorescent in situ hybridization (FISH)
Most sensitive technique for detecting and quantifying mRNA
Extremely small sample sizes can be used in the quantification of mRNA
Reverse transcription polymerase chain reaction (RT-PCR)
A solid surface to which a collection or microscopic DNA spots are attached
DNA microarray
Another name for DNA microarray
Biochip
DNA chip
Used to sequence the cDNAs corresponding to RNAs from the cells
RNA sequencing (RNA seq)
Used if the gene of interest has an unknown function
Inactivation of the gene
Another name for inactivation of the gene
Gene Knockout
A system that allows researchers to edit genes in living cells in a specific, desired way
CRISPR-Cas9
CRISPR-Cas9
Clustered regularly-interspaced short palindromic repeats
Advantages of using CRISPR-Cas9
Latest technology Faster Cheaper More accurate More efficient
Applications of DNA Technology
Medical Field DNA Forensics Manufacturing Industry Agriculture Environment
Altering the genes to treat or stop disease
Gene Therapy
