ASCP Molecular Biology Certification Exam with complete solutions.

ASCP Molecular Biology Certification Exam with complete solutions. Pyrimidine - Answer- One carbon ring Cytosine, Thymine, Uracil Purine - Answer- Two carbon rings Adenine, Guanine How are nucleotides joined together? - Answer- Condensation to form phosphodiester bond What is the function of mRNA? - Answer- Carries genetic info out of nucleus Transcript translated to protein What is the function of tRNA? - Answer- Carries aa to ribosome Anticodon pairs with codon on mRNA strand What is the function of rRNA? - Answer- part of ribosome structure most abundant RNA coordinated coupling of tRNA to mRNA codons Feedback inhibition - Answer- Product of pathway is noncompetitive inhibitor Binds to allosteric site to slow down rxn b/c too much product Exonucleases - Answer- Degrades nucleic acids by removing one terminal nt at a time Cleaves phosphodiester bond at end of chain 5' -- 3' and 3' -- 5' Endonucleases (Prok) - Answer- Restriction enzymes Cleaves phoshpodiester bonds w/i poly-nt chain Recognition site is palindromic sequence Types I-V ORI sites - Answer- nt sequence where replication is initiated Topoisomerase I - Answer- Induces ss breaks Remove DNA supercoils during TXN and DNA replication; for strand breakage during recombination; for chr condensation; and to disentangle intertwined DNA during mitosis topoisomerase II - Answer- cuts both strands of one DNA double helix, passes another unbroken DNA helix through it, and then reanneals the cut strands Gyrase (topoisomerase II) - Answer- Unwinds supercoiling caused by unwinding at the rep fork by introducing DSBs Helicase - Answer- Breaks H-bonds of double helix at the replication fork Primase - Answer- DNApol α (DNA dep RNA pol) adds short segments of complementary RNA to ssDNA template (primers), serves as starting points for replication single-strand DNA binding proteins (SSBPs) - Answer- Binds ssDNA and prevents it from re-annealing during TXN, replication, repair, and recombination Okazaki fragments - Answer- Short fragments of DNA synthesized by DNApol δ using the lagging strand (3'-5') as a template Ligase - Answer- Closes gaps in DNA Catalyzes phosphodiester bond between 3'OH and 5'P What are the steps in DNA replication? - Answer- 1. Initiate 2. Elongate 3. Terminate Telomeres - Answer- Repeat sequence (TTAGGG) at the ends of chr, protect chr from degradation RNA polymerase - Answer- DNA dependent RNApol Transcribes DNA template to RNA (3'--5'; anti-parallel) Splicesomes - Answer- Complex of snRNPs Removes introns from pre-mRNA and splices exons together Enhancers - Answer- Short regions of DNA that bind proteins (TXN factors) that enhance TXN of a gene Poly-A tail - Answer- Prevents mRNA from being degraded in cytoplasm 100-250 A's at 3' end 5' cap - Answer- 5'-5' pyrophosphate bridge to a methylated G added to 5' end of a mRNA Protects against degradation and as a recognition signal for TLN apparatus aminoacyl tRNA - Answer- tRNAs that carry amino acids Ribosomes - Answer- Where TLN occurs Prok: 30s and 50s Euk: 40s and 60s Catalyzes peptide bond between a.a.'s What is the path of a tRNA in a ribosome? - Answer- Acceptor Peptidyl Exit How is translation initiated? - Answer- small rRNA (40S) subunit binds mRNA and scans for start codon (AUG) Met-tRNA is brought to the P site Large rRNA (60S) subunit binds How is translation terminated? - Answer- Occurs when stop codon enters A site Release factor recognizes stop codon, hydrolyzes ester bond with P site, releasing aa chain Reverse transcriptase - Answer- enzyme that transcribes RNA to cDNA (lacks introns) RNA -- RNA:DNA -- cDNA (dsDNA) Pleiotrophy - Answer- a single gene controls the expression of many phenotypic traits ie Sickle Cell Anemia cDNA - Answer- intron free complementary DNA can be inserted into a plasmid Vector - Answer- helps carry DNA into cell ie plasmids, virus Open Reading Frame (ORF) - Answer- sections of DNA that begin with start codons and end with stop codons DNA: 5' -- 3' transcription: 3' -- 5' DNA -- RNA (promoter) translation: 5' -- 3' mRNA Spectrophotometer - Answer- Measures amount of light absorbed Quantitative measurement of [DNA/RNA] At what wavelength does DNA and RNA absorb? - Answer- 260 nm At what wavelength does protein absorb? - Answer- 280 nm Organic isolation method - Answer- 1. Lyse 2. Add phenol/ chloroform vortex/spin 3. Transfer aqueous layer (top) to new tube 4. Add chloroform:IAA (removes phenol) vortex/spin 5. Transfer aqueous layer to new tube 6. Add NaOAc and EtOH vortex/spin 7. Decant 8. Resuspend How do you inactivate RNases? - Answer- 200C for 2 hrs 30 min in 1M NaOH or quanidinum isothiocyanate Hybridization - Answer- 2 ssDNA molecules of comp base sequence can form a ds hybrid (duplex) What does the incubation step in hybridization do? - Answer- Allows formation of ds molecules Blocking DNA (Hybridization) - Answer- minimizes probe binding to nonspecific sequence ie salmon sperm DNA, Human LINE-1 Blocking Proteins (Hybridization) - Answer- minimize nonspecific binding of probe to membrane ie casein (milk), Denhardt's sol Stringency - Answer- conditions of hybridization that control the specificity of binding of the probe to the target sequence How can you increase strigency in a hybridization? - Answer- decrease [salt] increase [formamide] increase temp Formamide acts as a __________ in a hybridization. - Answer- denaturing agent Line Probe Assay (LiPA) - Answer- reverse hybridization assay using sequence-specific oligonucleotide probes (reverse SSOP) multi-parameter testing -- single strip Line Probe Assay steps - Answer- 1. Isolate nucleic acid (RNA) 2. Amplify 3. Hybridization 4. Strigent wash 5. Incubate with conjugate 6. Incubate with substrate 7. Detect .