ASCP Molecular Biology Certification
Exam
Pyrimidine correct answersOne carbon ring
Cytosine, Thymine, Uracil
What is the function of mRNA? correct answersCarries genetic info out of nucleus
Transcript translated to protein
What is the function of tRNA? correct answersCarries aa to ribosome
Anticodon pairs with codon on mRNA strand
What is the function of rRNA? correct answerspart of ribosome structure
most abundant RNA
coordinated coupling of tRNA to mRNA codons
Feedback inhibition correct answersProduct of pathway is noncompetitive inhibitor
Binds to allosteric site to slow down rxn b/c too much product
Endonucleases (Prok) correct answersRestriction enzymes
Cleaves phoshpodiester bonds w/i poly-nt chain
Recognition site is palindromic sequence
Types I-V
ORI sites correct answersnt sequence where replication is initiated
Topoisomerase I correct answersInduces 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 correct answerscuts both strands of one DNA double helix, passes another unbroken
DNA helix through it, and then reanneals the cut strands
Gyrase (topoisomerase II) correct answersUnwinds supercoiling caused by unwinding at the rep fork by
introducing DSBs
Helicase correct answersBreaks H-bonds of double helix at the replication fork
single-strand DNA binding proteins (SSBPs) correct answersBinds ssDNA and prevents it from re-
annealing during TXN, replication, repair, and recombination
Telomeres correct answersRepeat sequence (TTAGGG) at the ends of chr, protect chr from degradation
RNA polymerase correct answersDNA dependent RNApol
Transcribes DNA template to RNA (3'-->5'; anti-parallel)
Splicesomes correct answersComplex of snRNPs
Removes introns from pre-mRNA and splices exons together
Enhancers correct answersShort regions of DNA that bind proteins (TXN factors) that enhance TXN of a
gene
Reverse transcriptase correct answersenzyme that transcribes RNA to cDNA (lacks introns)
RNA --> RNA:DNA --> cDNA (dsDNA)
Open Reading Frame (ORF) correct answerssections 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 correct answersMeasures amount of light absorbed
Quantitative measurement of [DNA/RNA]
At what wavelength does DNA and RNA absorb? correct answers260 nm
At what wavelength does protein absorb? correct answers280 nm
Organic isolation method correct answers1. 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? correct answersAllows formation of ds molecules
Formamide acts as a __________ in a hybridization. correct answersdenaturing agent
Line Probe Assay (LiPA) correct answersreverse hybridization assay using sequence-specific
oligonucleotide probes (reverse SSOP)
multi-parameter testing --> single strip
Single-Stranded Conformational Polymorphism Ananlysis (SSCP) correct answersUsed 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
How does EtBr cause DNA to fluoresce? correct answersIntercalates into the double helix
Absorbs UV ~300 nm, emits ~600 nm
Pulse Field Gel Electrophoresis steps correct answers1. 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? correct answersDenature 90-96C
Anneal 50-70C
Extension 68-75C
Bisulfite DNA sequencing/Methylation specific correct answers1. 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 correct answers1. 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
Exam
Pyrimidine correct answersOne carbon ring
Cytosine, Thymine, Uracil
What is the function of mRNA? correct answersCarries genetic info out of nucleus
Transcript translated to protein
What is the function of tRNA? correct answersCarries aa to ribosome
Anticodon pairs with codon on mRNA strand
What is the function of rRNA? correct answerspart of ribosome structure
most abundant RNA
coordinated coupling of tRNA to mRNA codons
Feedback inhibition correct answersProduct of pathway is noncompetitive inhibitor
Binds to allosteric site to slow down rxn b/c too much product
Endonucleases (Prok) correct answersRestriction enzymes
Cleaves phoshpodiester bonds w/i poly-nt chain
Recognition site is palindromic sequence
Types I-V
ORI sites correct answersnt sequence where replication is initiated
Topoisomerase I correct answersInduces 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 correct answerscuts both strands of one DNA double helix, passes another unbroken
DNA helix through it, and then reanneals the cut strands
Gyrase (topoisomerase II) correct answersUnwinds supercoiling caused by unwinding at the rep fork by
introducing DSBs
Helicase correct answersBreaks H-bonds of double helix at the replication fork
single-strand DNA binding proteins (SSBPs) correct answersBinds ssDNA and prevents it from re-
annealing during TXN, replication, repair, and recombination
Telomeres correct answersRepeat sequence (TTAGGG) at the ends of chr, protect chr from degradation
RNA polymerase correct answersDNA dependent RNApol
Transcribes DNA template to RNA (3'-->5'; anti-parallel)
Splicesomes correct answersComplex of snRNPs
Removes introns from pre-mRNA and splices exons together
Enhancers correct answersShort regions of DNA that bind proteins (TXN factors) that enhance TXN of a
gene
Reverse transcriptase correct answersenzyme that transcribes RNA to cDNA (lacks introns)
RNA --> RNA:DNA --> cDNA (dsDNA)
Open Reading Frame (ORF) correct answerssections 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 correct answersMeasures amount of light absorbed
Quantitative measurement of [DNA/RNA]
At what wavelength does DNA and RNA absorb? correct answers260 nm
At what wavelength does protein absorb? correct answers280 nm
Organic isolation method correct answers1. 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? correct answersAllows formation of ds molecules
Formamide acts as a __________ in a hybridization. correct answersdenaturing agent
Line Probe Assay (LiPA) correct answersreverse hybridization assay using sequence-specific
oligonucleotide probes (reverse SSOP)
multi-parameter testing --> single strip
Single-Stranded Conformational Polymorphism Ananlysis (SSCP) correct answersUsed 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
How does EtBr cause DNA to fluoresce? correct answersIntercalates into the double helix
Absorbs UV ~300 nm, emits ~600 nm
Pulse Field Gel Electrophoresis steps correct answers1. 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? correct answersDenature 90-96C
Anneal 50-70C
Extension 68-75C
Bisulfite DNA sequencing/Methylation specific correct answers1. 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 correct answers1. 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
