Cell labb flashcards.txt
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• Which cells can be a source of DNA of certain gene?
All cells with a nucleus.
• Which cell can be a source of mRNA of certain gene?
Any cell which expresses that gene (synthesizes proteins from that gene.)
• What kinds of steps are in DNA isolation?
Cell lysis | Deproteination (removal of proteins) | (protease, absorption or extraction) | DNA precipitation by ethanol | DNA dilution in water or buffer
• Which methods are used for DNA isolation?
Phenol-chloroform extraction (different solubility conditions in solvents) | Salting out method (protein precipitation by NaCl) | Absorption method (silica-gel membrane)
• Which method is RNA detected by?
Northern blot. (side note; DNA detected by Southern blot.)
• How is measured a concentration of nucleic acids?
Spectrophotometry (absorbance maximum for nucleic acids 260 nm for proteins 280 nm)
• To which charge does DNA move towards in electrophoresis?
Positive, since DNA is negative itself.
• For which purpose is used gel electrophoresis?
To distinguish fragments of different sizes.
• Why is DNA seen in agarose gel?
Ethidium bromide is used as a dye for DNA (and RNA) and fluoresces under UV light.
• According to which constant does DNA migrate in gel electrophoresis?
Size of the DNA molecule or negative-to-positive migration???
• To which charge does DNA migrate in gel electrophoresis?
From negative to positive.
• What is it PCR?
Polymerase chain reation is a method to amplify DNA exponentially. Amplifies selected DNA sequences. DNA polymerase is used to replicate DNA by in vitro enzymatic replication.
• What is a mechanism of PCR machine?
A thermal cycler raises and lowers the temperature of the PCR samples in pre- programmed steps.
• Which 3 basic steps does one PCR cycle contain?
Denaturation, annealing, elongation (extension is the word used by Prof.Cerna)
• Which 3 basic components are added to PCR reaction?
DNA polymerase (Taq polymerase,) primers (forward & reverse,) and nucleotides. Also dNTPs???
• Which basic 4 components do you add to PCR reaction?
DNA strand, Primers, DNA Taq polymerase, dNTPs (Buffer + Cations(mg+))
• What is a mathematical formula of amplification run?
2 to the "n"th power = number of copies of wanted DNA region [where "n"=number of cycles]
• What is it hybridization?
It is the method used to detect the sequence of nucleotides of a DNA strand. It involves the denaturation of a DNA molecule, and the complementary binding of a "nucleic acid probe" to the original DNA strand (annealing.)
• What is it RFLP?
Restriction fragment length polymorphisms, or "riflips," are the differences in DNA sequence on homologous chromosomes that result in different patterns of restriction fragment lengths (restriction fragments=DNA segments resulting from treatment with restriction enzymes) useful as genetic markers. The restriction fragments are then seperated according to length by agarose gel electrophoresis.
• What is RFLP based upon?
Cut DNA into restriction fragments using suitable endonucleases.
• Which basic component is added to PCR product during RFLP?
Endonucleases (restriction enzyme)
• What do enzymes DNA polymerases produce and where are they used?
- DNA polymerases add nucleotides to an existing chain. The result is a complementary strand. Heat stable DNA polymerases such as Taq polymerase are used in PCR.
• What do restriction enzymes produce and where are they used?
Restriction enzymes (restriction endonuclease) produce restriction fragments. They are used in RFLP. They cut dsDNA at the sugar-phosphate backbone, leaving the nitrogenous base pairs (mostly) intact. They may however leave "sticky ends" consisting of a few bases without their complementary bases.
• What is a source of enzymes used in molecular biology?
• By which method is a complete gene analysis performed and its mechanism?
INSERT ANSWER HERE
• What is the method of finding unknown mutation in a gene?
• Which 3 methods are used for detecting on KNOWN mutation in a gene?
- PCR with allele specific primers (CFTR delta F508)
- PCR and RFLP (HFE C282Y)
- DNA hybridization with DNA probes
• What type of herebility is in disease cystic fibrose and hemachromostosis?
• What is the frequent mutation in cystic fibrose?
CFTR delta F508
• What is the frequent mutation in hemachromostasis?
• Which organs are impaired in c.f?
Lungs and digestive system.
• Which organs are impaired im hemachromostasis?
Liver, heart and pancreas
• What is it genomics and on what kind of method is it based?
Genomics is the study of an organism's entire genome.
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