DNA Polymerase in Lab vs. Cells Flashcards
DNA Polymerase in the cell has ________ timed replication of long chromosomes, ______ per cycle, and are done at _______ temperature.
precisely, once, same
Because synthesis is 5’ to 3’, replication of complementary DNA strands in the cell is ________ and is ______________.
asymmetric, semi-discontinuous
Many ____________ enzymes are required for fast high ________ replication of genomes such as helicase, DNA primase, and DNA ligase.
accessory, fidelity
In the lab, _______ _______ reaction amplifies short sequences of interest with steps controlled by ______
polymerase chain, temperature
DNA Polymerase active site has _______ , 3’ OH of ______ strand, nucleoside 5’triphosphate, 2Mg+.
template, growing
DNA ________ is the stepwise, directional addition of ______ to a growing DNA chain.
synthesis, dNTPs
DNA synthesis by polymerases require a _______ strand and a _______.
template, primer
DNA strands must be ________ prior to DNA synthesis.
separated
DNA polymerization always occurs in the ___ to ___ direction.
5’ , 3’
DNA double helix must be denatured for _________. In cells, denaturation is localized and starts at an ______ of replication.
replication, origin
A __________ _______ is where DNA replication in cells require many more components than the polymerase.
replication fork
The _________ of DNA strands is an ________-_________ process carried out by special enzymes.
separation, energy dependent
DNA helicases hydrolyze ____ to induce cyclic conformational changes and mechanical forces which are moving wedges that _____ the helix _____.
ATP, pry, apart
_____ _____ ______ proteins bind to the unwound strands of DNA to keep them separate and stop them from forming a helix again.
single strand binding
Because the DNA Double Helix is antiparallel, the synthesis of one DNA strand is __________ whereas it’s complementary strand is synthesized in ________.
continuous, fragments
DNA polymerases need an ____ primer. DNA primase is an ____ polymerase.
RNA, RNA
synthesis of the _______ strand leaves gaps that must be _______ by DNA _______.
lagging, sealed, ligase
DNA _______ seals gaps by charging the 5’ phosphate with ____ in the nick, which is subsequently attacked by 3’OH, sealing the DNA with phosphate.
ligase, AMP
_______ unwind DNA double helix ______ of the replication fork.
helicases, infront
IN PCR, the template is provided by ______ and can vary widely in ______.
you, quality
In PCR, DNA primers are provided by _______ and direct where synthesis ________.
you, starts
In PCR, the DNA strand is separated by ________ but a DNA polymerase that can _________ heat is required.
heating, withstand
PCR amplifies DNA through successive rounds of _________, _________, and __________ all regulated by temperature.
denaturation, annealing, extension
PCR results in the ___________ amplification of DNA molecules.
exponential
primer design requires finding the right _______ temperatures to balance specificity and structure.
melting
the melting temp (____) of a primer is the temp at which ___ of the DNA Duplex is denatured into single strand DNA. this reflects the _______ of the DNA sequence.
Tm, 1/2, stability
the Tm of primers depend on its ___________. It’s best predicted by _____ _________ calculations which account for basepair h bonds and base stacking stabilization.
sequence, nearest neighbor
_______ gel electrophoresis separates nucleic acids based on _____.
agarose, size
________ transcriptase allows for detection of RNA in addition to DNA
reverse
PCR can be used to amplify _________ DNA sequences from complex DNA mixtures.
specific
the power of PCR to single out specific DNA molecules can be exploited in the ______
clinic
Ex of Clinical use of PCR.
1. take blood sample from infected person
2. remove cells by _________
3. extract _____
4. _______ transcription and _____ amplification of particle
5. run sample through _____ _________ with column of uninfected person
centrifugation, RNA, reverse, PCR, gel electrophoresis
DNA tells what a cell ____ do, RNA tells what a cell ____ do.
can, does
reverse transcriptase is an RNA dependent DNA polymerase; it ______ RNA and ______ DNA
reads, writes
reverse transcriptase can be used to make ______ libraries. which are libraries of ________ after post transcriptional processing.
cDNA, mRNA’s
agarose gel electrophoresis doesnt require ______ as linear DNA molecules naturally have similar _____ and _____ proportional to their size.
SDS, shape, size
________ PCR directly detects the exponential growth of PCR products during the PCR in order to _____ specific DNA molecules. an example is the use of fluorophore to know when the strand has ______
quantitative, quantify, binded
in quantitative PCR a ________ ______ of known DNA quantities allows you to calculate the original concentration of DNA of known sequence from the ____.
standard curve, Ct
amount of DNA is _______ proportional to the cycle number.
inversely
________ sequencing is sequencing DNA using dideoxynucleotides (ddNTP) as chain __________. These have a 3’ H instead of OH.
sanger, terminators
____________ of nucleic acids facilitates easy detection of labeled probe sequences (florescent)
hybridization
microarray is an array of synthesized single stranded _____ corresponding to regions of a genome of interest
DNA
ChIP + detection
- cut DNA into _______
- immunoprecipitate ___ on a DNA binding protein
- detect bound ______
-detects where proteins bind
fragments
pull
DNA
