MCQ 4 Flashcards
Sanger sequencing
DNA sequencing based on selective incorporation of chain terminating dideoxynucleotides by DNA polymerase during in-vitro replication.
How do dideoxynucleotides (ddNTPS) function?
Lacking a hydroxyl group they prevent further extension of a DNA strand, resulting in fragments of different lengths.
Role of dNTPs in sanger sequencing.
Standard nucleotides that DNA polymerase incorporate into the growing DNA strand providing a substrate for elongation.
Primers in sanger sequencing
Short sequences that provide a starting point for DNA synthesis during sanger sequencing.
Visualization after sanger sequencing
ddNTP is labeled with differing fluorescent dyes, allowing the fragments to be visualized and realized in gel electrophoresis.
Sanger vs PCR sequencing
Sanger sequencing typically uses a single primer, PCR uses a pair of primers.
Recombinant gene vectors
A DNA molecule used as a tool to artificially introduce a foreign piece of DNA into another cell to be replicated or expressed.
OriV
Stands for origin of replication is a sequence within a plasmid that allows it to autonomously replicate inside host cells independent of the host’s chromosomal DNA replication.
Antibiotic resistance testing.
Antibiotic resistant genes are included in plasmids to allow researchers to select for cells that have successfully taken up the plasmid.
Bacteriophage in recombinant technology
Phages can be engineered to be used as vectors to deliver genetic information.
Advantages of using yeast artificial chromosomes (YACs)
Can clone larger fragments than bacterial chromosomes.
Cosmid vs fosmid vectors
Fosmids are based on f-plasmids and are more stable, reducing the likelihood of chimeric inserts.
Oligonucleotides
Short DNA fragments used as primers in sequencing, normally to define starting points.
Temperature in PCR vs Sanger
PCR requires cyclical changes in temperature, sanger runs at constant temperatures.
Importance of LacZ gene
Reporter in cloning vectors to identify successful insertions via blue/white screening.
Multiple cloning sites (MCSs)
Short DNA sequence in plasmid vector that contain restriction sites for several types of restriction enzymes.
