M1L3 Prt 2 200 QS

Nucleotides

What are the basic monomer building blocks that make up DNA?

Adenine and Guanine

Which nitrogenous bases are classified as larger, double-ring structures called purines?

Cytosine and Thymine

Which nitrogenous bases are classified as smaller, single-ring structures called pyrimidines in DNA?

Adenine

Which specific purine pairs with Thymine in DNA?

Guanine

Which specific purine pairs with Cytosine in DNA?

Deoxyribose

What five-carbon pentose sugar is found in DNA that provides structural stability?

Hydroxyl group (-OH)

What specific chemical group is missing at the 2nd prime carbon position in deoxyribose compared to ribose?

Phosphodiester bond

Which chemical bond connects the 3' carbon of one pentose sugar to the 5' carbon of another sugar in a nucleic acid strand?

Phosphate group

What component of the nucleotide imparts an acidic character and a net negative charge to DNA?

Nucleoside

What is a molecule consisting solely of a nitrogenous base and a pentose sugar (Base + Sugar) called?

Deoxynucleoside triphosphates (dNTPs)

What building blocks are used by DNA polymerases during PCR and sequencing reactions?

James Watson & Francis Crick

Who proposed the famous double helix structure of DNA in 1953?

Rosalind Franklin

Whose X-ray diffraction data was crucial for establishing the classic DNA double helix model?

2

How many hydrogen bonds connect Adenine and Thymine in DNA?

3

How many hydrogen bonds connect Guanine and Cytosine in DNA?

Antiparallel

What term describes the opposite orientation of two DNA strands running in 5' to 3' and 3' to 5' directions?

Leading strand

What do you call the DNA strand that is synthesized continuously during replication?

Lagging strand

What do you call the DNA strand that is synthesized discontinuously during replication?

Major and Minor grooves

Which DNA structural feature allows transcription factors and proteins to access base sequences without unwinding the double helix?

Chargaff's Rules

What rule states that in double-stranded DNA, the percentage of adenine equals thymine and the percentage of guanine equals cytosine?

B-DNA

What is the standard, biologically active, right-handed helix form of DNA under physiologic cellular conditions?

Right-handed

What is the helix direction of A-DNA?

~10.5

How many base pairs per turn are present in B-DNA?

Z-DNA

Which form of DNA is a left-handed helix with approximately 12 base pairs per turn, occurring under GC-rich sequences or torsional stress?

260 nm

At what wavelength in nanometers do nucleic acids/DNA absorb UV light maximally due to their aromatic nitrogenous bases?

280 nm

At what wavelength in nanometers do proteins absorb UV light maximally due to aromatic amino acids like tryptophan and tyrosine?

Approximately 1.8

What is the expected A260/A280 absorbance ratio for pure DNA?

Protein

An A260/A280 ratio of 1.6 or lower indicates contamination by what molecule?

RNA

An A260/A280 ratio greater than 2.0 indicates potential contamination by what molecule?

Guanidine / Phenol

Absorbance at 230 nm assesses contamination by organic compounds, salts, or residual reagents such as which chemical?

2.0 to 2.2

What is the ideal A260/A230 ratio for nucleic acid purity?

Nanodrop

What microvolume UV-Vis spectrophotometer requires only 1 to 2 µL of sample for quantification?

It cannot distinguish DNA from RNA or single-stranded DNA

What is a key limitation of UV spectrophotometry via Nanodrop?

PicoGreen (or SYBR Green I)

Name a double-stranded DNA-intercalating fluorescent dye used in fluorometric quantification methods that is more sensitive than UV-Vis.

Positive electrode (Anode)

In agarose gel electrophoresis, toward which charged electrode does DNA migrate?

Smearing

What visual indication on an agarose gel suggests DNA degradation rather than intact DNA?

Uracil

Which pyrimidine base replaces Thymine in RNA?

It possesses a hydroxyl (-OH) group at the 2' carbon position of ribose

Why is RNA inherently less stable and more prone to hydrolysis than DNA?

Ribozymes

What type of RNA serves as catalytic RNA behaving like an enzyme?

mRNA (Messenger RNA)

What type of RNA carries the genetic blueprint from DNA to the ribosome to be translated into protein?

rRNA (Ribosomal RNA)

Which type of RNA constitutes approximately 80% of total cellular RNA in prokaryotes like E. coli?

Cloverleaf shape

What is the secondary structure shape of a transfer RNA (tRNA) molecule?

L-shape

What is the tertiary structure shape of a transfer RNA (tRNA) molecule?

snRNA (Small nuclear RNA)

Which small regulatory RNA is responsible for removing introns from pre-mRNA during splicing?

snoRNA (Small nucleolar RNA)

Which small regulatory RNA processes and chemically modifies rRNA in the nucleolus?

microRNA (miRNA) / siRNA (Small interfering RNA)

Which small regulatory RNAs regulate gene expression by blocking translation or promoting mRNA degradation?

RNA polymerase

What enzyme performs DNA-directed RNA synthesis during transcription?

Template strand (or Antisense strand)

During transcription, what is the DNA strand called that is actually read by RNA polymerase to make RNA?

Gene

A linear sequence of DNA that carries instructions to produce a functional RNA or protein is called a what?

Locus (plural: Loci)

What Latin-derived term refers to a specific place or region of interest within a gene?

rpoB gene

What clinically significant gene in Mycobacterium tuberculosis encodes the RNA polymerase beta subunit and has mutations linked to rifampicin resistance?

N gene (Nucleocapsid protein gene)

In SARS-CoV-2 molecular detection, what highly expressed gene is commonly targeted as a diagnostic locus?

Flanking

What term describes primer binding to sequences on either side of a target diagnostic locus in PCR?

Codon

A combination of three consecutive nucleotides within a gene that specifies a particular amino acid is called a what?

ATG (in DNA) / AUG (in RNA)

What universal nucleotide triplet serves as the start codon and specifies methionine?

3' to 5' direction

In what direction is the DNA template strand read by RNA polymerase during transcription?

5' to 3' direction

In what direction does the synthesized RNA strand grow during transcription?

One

How many types of RNA polymerase exist in prokaryotes like E. coli?

Holoenzyme

What complex consists of the prokaryotic core RNA polymerase combined with the sigma factor?

Sigma factor (σ)

Which subunit of prokaryotic RNA polymerase recognizes the promoter region and guides the enzyme to the start site?

Alpha (α) subunits (2 copies)

Which structural subunits assemble the prokaryotic RNA polymerase complex and bind regulatory proteins?

Beta (β) subunit

Which subunit of prokaryotic RNA polymerase contains the active site for phosphodiester bond formation?

Beta prime (β') subunit

Which subunit of prokaryotic RNA polymerase binds and holds the DNA template strand during transcription?

Omega (ω) subunit

Which subunit stabilizes the overall prokaryotic RNA polymerase complex and assists in assembly?

RNA Polymerase I

Located in the nucleolus, which eukaryotic RNA polymerase synthesizes most rRNA molecules?

RNA Polymerase II

Located in the nucleoplasm, which eukaryotic RNA polymerase synthesizes mRNA, snRNA, and microRNA?

RNA Polymerase III

Located in the nucleoplasm, which eukaryotic RNA polymerase synthesizes tRNA and 5S rRNA?

TATA box

What conserved promoter sequence motif is typically recognized by RNA polymerase II in eukaryotes?

-10 and -35 regions

What regions within prokaryotic promoters are recognized by the RNA polymerase holoenzyme?

ATP, GTP, CTP, UTP (NTPs)

What are the four nucleotide substrates (triphosphates) required for RNA synthesis?

Transcription bubble

What unwound DNA structure is created when RNA polymerase binds to the promoter and separates the DNA strands?

It lacks proofreading capability

Why is RNA polymerase more error-prone than DNA polymerase during elongation?

Rho factor (Rho protein)

In prokaryotic transcription termination, what protein binds to the nascent RNA and chases RNA polymerase to cause detachment?

GC-rich hairpin loop followed by a run of uracils

What structural feature formed by the RNA transcript is essential for Rho-independent transcription termination?

5' Cap (7-methylguanosine cap)

What modified nucleotide structure is added to the 5' end of eukaryotic pre-mRNA shortly after transcription begins?

Protects RNA from exonuclease degradation, facilitates nuclear export, and helps ribosome recognition

What is the main function of the 5' cap on mRNA?

3' Polyadenylation (Poly-A tail)

What modification involves adding ~200 adenine nucleotides to the 3' end of eukaryotic mRNA?

Exons

What are the coding sequences of a eukaryotic pre-mRNA called?

Introns

What are the non-coding intervening sequences of pre-mRNA called that must be removed?

Spliceosome

What multi-megadalton complex made of snRNAs and proteins catalyzes pre-mRNA splicing?

Alternative splicing

What process allows a single eukaryotic pre-mRNA gene to produce multiple protein isoforms by joining different combinations of exons?

RNA editing

What post-transcriptional process chemically alters an RNA sequence post-synthesis by inserting, deleting, or substituting bases?

RNases (Ribonucleases)

What ubiquitous, highly resilient enzymes pose the greatest threat to RNA sample integrity in laboratory settings?

DEPC (Diethyl pyrocarbonate) / RNase Zap

What chemical treatment agent is commonly used to treat water and glassware to deactivate RNases?

Guanidinium thiocyanate

What strong chaotropic agent in cell lysis buffers denatures proteins and inhibits RNase activity during RNA extraction?

TRIzol (Guanidinium thiocyanate-phenol-chloroform)

What reagent mixture is used in classical organic phase separation of RNA, DNA, and proteins?

Aqueous phase (top layer)

Following chloroform addition and centrifugation in a TRIzol extraction, in which layer/phase is pure RNA found?

Interphase (DNA) and Organic phase (Proteins/Lipids)

In a TRIzol extraction phase separation, where are DNA and proteins located, respectively?

Silica membrane

In column-based RNA extraction, to what solid-phase material does RNA selectively bind under high salt conditions?

Oligo (dT) columns

What matrix component is utilized in affinity columns to specifically hybridize and isolate mRNA based on its poly-A tail?

rRNA constitutes over 80% of total cellular RNA and can obscure low-abundance transcripts

Why is rRNA frequently depleted before RNA sequencing or transcriptomic studies?

RNase H

What enzyme selectively degrades RNA in RNA-DNA hybrids during specific rRNA depletion protocols?

40 µg/mL

An A260 reading of 1.0 in UV spectrophotometry corresponds to approximately how much concentration of RNA?

28S and 18S rRNA

Which two prominent ribosomal RNA bands are evaluated on an agarose gel to determine eukaryotic RNA integrity?

2:1 ratio

What ratio between 28S and 18S rRNA intensity indicates high-quality, intact eukaryotic RNA?

RNA Integrity Number (RIN)

What metric scaled from 1 (degraded) to 10 (intact) is generated by automated systems like the Agilent Bioanalyzer to grade RNA quality?

Reverse transcription

What process converts RNA into complementary DNA (cDNA)?

Reverse transcriptase

What viral enzyme synthesizes single-stranded cDNA using an RNA template?

M-MLV (Moloney Murine Leukemia Virus)

Name a common reverse transcriptase source derived from a murine retrovirus.

AMV (Avian Myeloblastosis Virus)

Name a highly thermostable reverse transcriptase source used when RNA has extensive secondary structures.