ASCP Molecular Biology Certification Exam 10 studiers today Science Biology Pathology 4.8 (11 reviews) Students also viewed Terms in this set (1434) MB (ASCP) 2020 Teacher 402 terms Kquan20 Preview Molecular Diagnostics Chapter 2 Teacher 57 terms Lolo20000000 Preview Membranes 29 terms Pyrimidine One carbon ring Cytosine, Thymine, Uracil What is the function of mRNA? Carries genetic info out of nucleus Transcript translated to protein What is the function of tRNA? Carries aa to ribosome Anticodon pairs with codon on mRNA strand What is the function of rRNA? part of ribosome structure most abundant RNA coordinated coupling of tRNA to mRNA codons Feedback inhibition Product of pathway is noncompetitive inhibitor Binds to allosteric site to slow down rxn b/c too much product ASCP Molecular Biology Certification Exam Flashcards | Quizlet https://quizlet.com/255480411/ascp-molecular-biology-certification-exa... 1 of 11 10/06/2024, 21:03 Endonucleases (Prok) Restriction enzymes Cleaves phoshpodiester bonds w/i poly-nt chain Recognition site is palindromic sequence Types I-V ORI sites nt sequence where replication is initiated Topoisomerase I 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 cuts both strands of one DNA double helix, passes another unbroken DNA helix through it, and then reanneals the cut strands Gyrase (topoisomerase II) Unwinds supercoiling caused by unwinding at the rep fork by introducing DSBs Helicase Breaks H-bonds of double helix at the replication fork single-strand DNA binding proteins (SSBPs) Binds ssDNA and prevents it from re-annealing during TXN, replication, repair, and recombination Telomeres Repeat sequence (TTAGGG) at the ends of chr, protect chr from degradation RNA polymerase DNA dependent RNApol Transcribes DNA template to RNA (3'-->5'; antiparallel) Splicesomes Complex of snRNPs Removes introns from pre-mRNA and splices exons together Enhancers Short regions of DNA that bind proteins (TXN factors) that enhance TXN of a gene Reverse transcriptase enzyme that transcribes RNA to cDNA (lacks introns) RNA --> RNA:DNA --> cDNA (dsDNA) ASCP Molecular Biology Certification Exam Flashcards | Quizlet https://quizlet.com/255480411/ascp-molecular-biology-certification-exa... 2 of 11 10/06/2024, 21:03 Open Reading Frame (ORF) 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 Measures amount of light absorbed Quantitative measurement of [DNA/RNA] At what wavelength does DNA and RNA absorb? 260 nm At what wavelength does protein absorb? 280 nm Organic isolation method 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 What does the incubation step in hybridization do? Allows formation of ds molecules Formamide acts as a __________ in a hybridization. denaturing agent Line Probe Assay (LiPA) reverse hybridization assay using sequence-specific oligonucleotide probes (reverse SSOP) multi-parameter testing --> single strip Single-Stranded Conformational Polymorphism Ananlysis (SSCP) Used for known gene, unknown mut Mutation screening Short PCR products form 3D conformation when cooled --> muts have different conformation than WT Non-denaturing PAGE, muts migrate different than WT ASCP Molecular Biology Certification Exam Flashcards | Quizlet https://quizlet.com/255480411/ascp-molecular-biology-certification-exa... 3 of 11 10/06/2024, 21:03 How does EtBr cause DNA to fluoresce? Intercalates into the double helix Absorbs UV ~300 nm, emits ~600 nm Pulse Field Gel Electrophoresis steps 1. culture 2. embed pellet in agarose plug 3. treat w/ lysozyme (cell lysis) 4. proteinase K 5. gel What are the 3 steps of PCR and their temperatures? Denature 90-96C Anneal 50-70C Extension 68-75C Bisulfite DNA sequencing/Methylation specific 1. RE digest 2. Electrophorese and purify fragment of interest 3. Denature and incubate w/ sodium bisulfate (turns C>U, methylated C is unchanged) 4. clean, ppt, and resuspend 5. PCR --> sequence 6. Compare treated vs untreated, note where CG are not changed to TA NASBA steps 1. Hybridize oligo-T7P primer to target seq 2. RT/RNase H 3. Hybridize with target-specific oligo primer (P2) 4. RNA transcript of T7 RNA pol Do you have to know the gene sequence in order to do DNA sequencing? Yes, in order to design primers You do NOT need to know the mutation Sanger sequencing method Divided into 4 samples (ddA, T, G, C) Label with radioactive/dye oligo at 3' end Mix with taq, dNTPs, ddNTP and incubate run on gel --> frags will terminate at different lengths Fluorescent in situ hybridization (FISH) Uses fluorescent probes to detect DNA sequences on chr ASCP Molecular Biology Certification Exam Flashcards | Quizlet https://quizlet.com/255480411/ascp-molecular-biology-certification-exa... 4 of 11 10/06/2024, 21:03 What types of probes are used for FISH? Dual fusion: 2 probes flank the breakpoint at both t locations CEN probes: centromeric probes bind to repetitive alpha satellite sequences Telomeric probes Whole chr paints What is the wavelength for background in spectrophotometery? 320 nm How do you determine quality of DNA/RNA using a spectrophotometer? A260/A280 What is considered good quality DNA/RNA from spectrophotometry? DNA: 1.7-2.0 RNA: 2.0-2.3 What is considered poor quality RNA/DNA from spectrophotometry? <1>