PCR & DNA TECH
What are the three main steps of PCR that repeat in each cycle?
Denaturation (heating to separate strands)
Annealing (cooling so primers attach)
Extension (DNA polymerase builds complementary strands)
What percentage of the human genome actually codes for proteins (exons)?
About 2.5%
What does CRISPR technology allow scientists to do?
Edit DNA by making targeted, precise changes to specific nucleotide sequences in genes
What modification makes Sanger sequencing work, and what happens when these are added?
Di-deoxynucleotides (modified nucleotides) that terminate the DNA chain when added, stopping synthesis
Both plant and animal cells have mitochondria. True or False: Only animal cells can perform cellular respiration.
False - both plant and animal cells have mitochondria and perform cellular respiration to produce ATP
If you run PCR for 10 cycles starting with 1 DNA molecule, how many copies will you have?
1,024 copies (2^10 = 1,024)
What is chromatin made of?
DNA wrapped around histone proteins (DNA-protein complex)
Who won the 2020 Nobel Prize in Chemistry for inventing CRISPR technology?
Jennifer Doudna and Emmanuelle Charpentier
In Sanger sequencing, how are the four different di-deoxynucleotides (diA, diC, diG, diT) identified?
Each is tagged with a different fluorescent color that glows when detected
What is the difference between primary and secondary active transport in terms of energy use?
Primary active transport uses ATP directly to move substances against their gradient. Secondary active transport uses the energy stored in concentration gradients (created by primary transport) to move other substances - it doesn't use ATP directly
Why must the primers in PCR have their 3' ends oriented toward each other?
Because DNA polymerase can only add nucleotides to the 3' end, so the primers need to point toward each other for the DNA to be copied in both directions
What is a nucleosome and what does it consist of?
A nucleosome is the basic unit of DNA packaging, consisting of 8 histone proteins wrapped with about 150 base pairs of DNA
What are restriction enzymes used for, and what do they recognize?
Restriction enzymes cut DNA at specific recognition sequences (specific base pair sequences) on double-stranded DNA - used for recombinant DNA and genome editing
What does "shotgun sequencing" mean for whole genome sequencing?
The entire genome is broken into many small, overlapping fragments, each fragment is sequenced separately, then the sequences are assembled by aligning overlapping regions
If you have an mRNA sequence 5'-AUGCGUUAA-3', what would be the anticodon sequence for the tRNA that brings in the second amino acid?
3'-GCA-5' (the second codon is CGU, so the anticodon is 3'-GCA-5', antiparallel to the codon)
In gel electrophoresis, do smaller or larger DNA fragments travel farther through the gel, and why?
Smaller fragments travel farther because they can move more easily through the pores in the gel matrix
What are transposons, and what percentage of the human genome do they make up?
Transposons are "jumping genes" - DNA sequences that can copy themselves and move to other locations in the genome. They make up about 45% of the human genome
What's the difference between "sticky ends" and "blunt ends" when restriction enzymes cut DNA?
Sticky ends have overhangs (uneven cuts) with unpaired bases that can base-pair with complementary sequences, while blunt ends are even cuts with no overhangs on both strands
What advantage do "Next-Gen" sequencing methods have over Sanger sequencing?
Next-gen methods sequence many fragments simultaneously (higher throughput), are faster, more automated, and sequence as they synthesize (real-time fluorescence), making analysis more rapid
Name three things that can make cell membranes MORE fluid.
Higher temperature, shorter fatty acid chains, more unsaturated fatty acids (with double bonds), or adding cholesterol at LOW temperatures
What special property must the DNA polymerase used in PCR have that normal DNA polymerase doesn't need?
It must be heat-stable (thermostable) because PCR involves repeated heating to 95°C for denaturation, which would denature normal polymerase
Why doesn't genome size or gene number reliably predict biological complexity in eukaryotes?
Because organisms with larger genomes don't necessarily have more genes or greater complexity - much of the genome is non-coding DNA, and complexity depends more on gene regulation and protein diversity than sheer number of genes
Explain how recombinant DNA is made using restriction enzymes and what enzyme seals the final product.
Restriction enzymes cut both the donor DNA fragment and the vector (like a plasmid) at recognition sites, creating complementary sticky ends. The fragments are mixed and their sticky ends base-pair together, then DNA ligase seals the sugar-phosphate backbone to create the final recombinant DNA molecule
In next-gen sequencing, how does the system detect which nucleotide has been added to the growing strand?
When a complementary nucleotide is added, pyrophosphate (PPi) is released, which causes a flash of light that is recorded. No flash means that nucleotide wasn't complementary and wasn't added
What is genome annotation used for? A) Determining the size of a genome B) Identifying and labeling types of sequences like genes, regulatory elements, and repetitive DNA C) Comparing genomes between different species only D) Packaging DNA into chromosomes
B - Genome annotation identifies and labels different types of sequences (genes, open reading frames, tandem repeats, transcription factor binding sites, introns/exons) and their locations in the genome