Molecular Diagnostics Chapter 4 Actual Study Questions with Correct Answers

Molecular Diagnostics Chapter 4 Actual Study Questions with Correct Answers 1. Which whole blood fraction is the most abundant source of genomic DNA? - ANSWER Buffy Coat 2. Which specimen will yield the best DNA preparation? - ANSWER Fresh Blood 3. agarose gel - ANSWER a jelly-like slab used to separate molecules on the basis of molecular weight, polysaccharide that comes from seaweed 4. concentration of agarose in a gel determines the size of spaces in the gel. 3% agarose resolution is best for _______bp of DNA. 0.6% is best ________bp of DNA. - ANSWER 50-100 bp and 1000, 20,000 5. Are low concentration agarose gels easily broken? - ANSWER yes 6. Electro endosmosis - ANSWER This electrophoresis error is caused by cations in the buffer moving in the opposite direction which slows/distorts migration of samples 7. Pulse field gel electrophoresis (PFGE) - ANSWER used to separate especially long strands of DNA by length in order to tell differences among samples. Does so by alternating current in different dimensions during migration of sample. Requires long separation times and temperature control. 50,000-250,000bp 8. FIGE - ANSWER field-inversion electrophoresis, samples move forward and backward due to the changing current/pulse field 9. polyacrylamide gel - ANSWER -porous matrix mixture used in electrophoresis typically for smaller fragments -single bp resolution so can be used for sequencing and mutation analysis -synthetic can be manipulated to have exact same conditions or necessary conditions 10.w/v, t, C PAGE gels IF, T value 6% and 19:1 acrylamide:bis, then C=? - ANSWER w/v total percentage concentration of acrylamide and t the percentage of monomer, T=acrylamide and cross linker, C= percentage of monomer that is a crosslinker. C=1/20 or 5% 11.Can PAGE gels polymerize upon cooling? - ANSWER No need catalyst such as APS and TEMED or light 12.How is RNA denatured during electrophoresis? - ANSWER more challenging because secondary structure bonding is stronger compared to DNA hybridization between complementary strands, denaturant MMH, formaldehyde, or glyoxal can be used 13.What is the advantage of SYBR green over EtBr for gel electrophoresis nucleic acid staining? - ANSWER EtBr is a mutagen and is toxic, SYBR green is not 14.Silver staining - ANSWER used to detect nucleic acids after gel electrophoresis, gel is washed with silver nitrate or silver diamine and stained the precipitated with formaldehyde 15.Which fixative is the least damaging to nucleic acids? - ANSWER Buffered formalin 16.The most abundant form of RNA in all cells is what? - ANSWER Ribosomal 17.Diethyl pyrocarbonate (DEPC) is a chemical that is used to do what? - ANSWER Inactivate RNAase 18.What will not inhibit RNAses? - ANSWER Tris buffers 19.In organic isolation of RNA procedures, guanidinium isothiocyanate is added to what? - ANSWER Inhibit RNAse's 20.PolyT oligomers bound to a matrix resin column will selectively isolate what? - ANSWER Messenger RNA 21.Nucleic acid concentration can be assessed relatively simply and quickly by using what procedures? - ANSWER Electorphoresis 22.What is the concentration of DNA whereby a 1:100 dilution has an absorbance reading of 0.020 at 260 nm? - ANSWER 0.020 x 50 x 100 = 100ug/ml