Ch 20 Gen Coding
What is a plasmid?
A small circular DNA molecule often used as a vector in gene cloning (Vectors are commonly this) (Naturally occurring plasmids that carry confer resistance to antibiotics are called R Factors, most modern plasmids are derived from these)
When cloning a gene into a vector, the sugar-phosphate backbone of each DNA molecule is covalently linked by the enzyme
DNA Ligase
Order the steps in one cycle of a PCR reaction, putting the first step at the top.
Denaturation- Strands separate (due to heat) Primer annealing- Primers bind to the DNA (When temp is lowered) Primer extension- Doubles the amount of the template DNA (Heated up again) starting at the primers in the 5' to 3' direction.
The DNA sequencing method developed by Frederick Sanger that became a commonly used method of DNA sequencing is called ______ sequencing.
Dideoxy sequencing
Northern Blots
Identify specific RNA within a mixture (where and how much)
The use of in vitro molecular techniques that combine DNA fragments to produce novel arrangements is called _________ DNA technology .
Recombinant (Used to manipulate fragments of DNA to produce new arrangements)
A molecule that has covalently linked DNA fragments from at least two sources is called a(n) molecule.
Recombinant DNA
Enzymes that bind to a specific DNA sequence and cut the DNA backbone are called
Restriction endonucleases
Chromosomal DNA is a common source of cloned DNA.
TRUE
In PCR, why do the primers bind to specific sites in the DNA on either side of the gene of interest?
They are complementary to the flanking sequences.
In dideoxy sequencing, if a dideoxyribonucleotide is incorporated into the growing strand of DNA, the strand can no longer grow as there is no 3' OH group. This is called ______.
chain termination
To make many copies of a gene, you would _________ that gene .
clone or amplify - used to find/ study a specific gene or proteins and its function or structure.
Transformation occurs when ______.
competent cells take up DNA from the medium
Restriction endonucleases are used in gene cloning to ______.
cut the DNA backbone prior to inserting the DNA to be cloned
If the oxygens on carbons 2 and 3 of the sugar of a nucleotide have been removed, the nucleotide is referred to as a
dideoxyribonucleotides
The replication of recombinant DNA molecules inside a host cell is one form of ______.
gene Coding
Select the reagents needed to make cDNA.
mRNA (mixed with primer) Poly-dT primer( short strand of dna complementary to 3' mRNA) dNTPs Reverse transcriptase (both added to complementary end of mRNA (made via reverse transcriptase, reverse transcriptase uses RNA as a template to make double stranded DNA molecule)
The enzyme that uses RNA as a template to make a complementary strand of DNA is called
reverse transcriptase
In PCR, the DNA to be amplified is called the
template DNA
The process by which competent cells take up DNA from the extracellular medium is called
transformation (following ligation)
In dideoxyribonucleotides, ___________ oxygens are removed from the sugar compared with ribose.
two (ddNTPs, removal makes is so the DNA strand can no longer grow, this is called Chain termination)
Why is Taq polymerase used in PCR?
(Stable at high temperatures) The DNA polymerase must be thermostable as PCR involves cycles of heating.
Western Blots
*identify protein molecules on a gel.* -involves separating proteins by gel electrophoresis and *probing with labeled antibodies* for specific proteins
Order the following steps in cloning a gene, putting the first step at the top.
-Chromosomal DNA is isolated and cut with a restriction enzyme; the plasmid DNA is cut with the same enzyme -The digested chromosomal DNA and plasmid DNA are incubated together -Ligation by DNA ligase
In gene cloning, what is the vector?
A small DNA molecule that can replicate independently within a host cell (Acts as the carrier of the DNA segment that we want to clone)(Most are plasmids some are viruses, if plasminds they are called R Factors)