Genetics Lab Presentation Questions

What are some good characteristics of a primer?

A good primer is about 18–25 base pairs long, specific to the target sequence, and has a balanced GC content (40–60%)

What are primers used for?

Primers are used to start DNA synthesis by providing a starting point for DNA polymerase during PCR

How is a primer used to identify a gene in PCR?

A primer identifies a gene by binding specifically to sequences on either side of the target gene so that only that gene is amplified

What could happen if my primer pairs are less than 20 bp long?

If primers are less than 20 base pairs long, they may bind nonspecifically to other DNA sequences and produce incorrect PCR products.

Which temperature in PCR is important for primers?

The annealing temperature is important because it determines how well the primers bind to the target DNA.

Why is it important to extract DNA?

It is important to extract DNA so that it can be studied, analyzed, or used in experiments such as PCR or sequencing.

What does RNase do?

RNase is an enzyme that breaks down RNA so that only DNA remains in the sample.

How does a centrifuge work?

A centrifuge works by spinning samples at high speed to separate components based on their density.

Why did we use a centrifuge to isolate DNA?

We used a centrifuge to separate DNA from other cell components by forcing heavier materials to form a pellet at the bottom.

If we are to isolate DNA from human cells instead of bacteria, how might the process be different?

The process may require stronger lysis steps and removal of more proteins because human cells contain more complex structures than bacteria.

What is the purpose of PCR?

The purpose of PCR is to amplify a specific DNA sequence to produce many copies for analysis.

What is one difference between PCR and DNA replication?

PCR occurs in a laboratory using temperature changes, while DNA replication occurs naturally inside cells using enzymes.

Why is PCR important?

PCR is important because it allows scientists to detect, identify, and study small amounts of DNA quickly.

What are the three main steps of PCR, and what happens at each step?

The three steps are denaturation (DNA strands separate), annealing (primers bind to DNA), and extension (DNA polymerase builds new DNA strands).

What was the purpose of running PCR on the four bacteria?

The purpose was to amplify a specific gene from each bacterium so the DNA could be analyzed and compared.

What properties of DNA help it move through a gel?

DNA moves through a gel because it has a negative charge and small size, allowing it to migrate toward the positive electrode.

What happens if a smear appears on a gel?

A smear on a gel usually indicates degraded DNA, contamination, or nonspecific amplification.

What does a faint band at the bottom of a gel represent?

A faint band at the bottom of a gel usually represents very small DNA fragments or primer dimers.

Why is a DNA ladder needed?

A DNA ladder is needed to estimate the size of DNA fragments by comparing them to known fragment lengths

What allows us to visualize DNA bands on a gel?

DNA bands are visualized using a DNA-binding stain such as ethidium bromide or SYBR Safe under UV light.

What is the loading dye used for?

Loading dye is used to make the sample visible and help it sink into the wells of the gel.

Why is it important to purify PCR product?

It is important to purify PCR product to remove primers, enzymes, and contaminants before further analysis like sequencing.

What is the importance of a spin column?

A spin column is important because it binds DNA while allowing unwanted substances to be washed away.

What technique(s) are used to purify PCR product?

PCR product is purified using centrifugation with a spin column and washing buffers.

How could you check for contamination in your PCR product?

You can check for contamination by running a negative control sample on a gel to see if unexpected DNA bands appear.

What is one major difference between Maxam-Gilbert and Sanger sequencing?

Maxam-Gilbert sequencing uses chemical cleavage of DNA, while Sanger sequencing uses chain-terminating nucleotides during DNA synthesis.

What is shotgun sequencing?

Shotgun sequencing is a method where DNA is randomly broken into small fragments and then sequenced and assembled into a complete sequence.

What components are needed to perform a sequencing reaction?

A sequencing reaction requires DNA template, primer, DNA polymerase, nucleotides, and labeled chain-terminating nucleotides.

What is the result of a sequencing reaction?

The result of a sequencing reaction is the exact order of nucleotides in a DNA sequence.

What is bioinformatics?

Bioinformatics is the use of computer programs to analyze and interpret biological data such as DNA sequences.

What major programs were used to analyze the ctrA gene? What did each one do?

Programs like BLAST identified similar sequences, and sequence alignment tools compared DNA or protein sequences to determine gene function.

What is the difference between BLASTN and BLASTX?

BLASTN compares DNA sequences to DNA, while BLASTX compares DNA sequences translated into protein to protein databases.

What is the composition of a promoter in bacteria?

A bacterial promoter typically contains a -10 region (TATAAT) and a -35 region that help RNA polymerase bind and start transcription.

Why are there only 20 amino acids, but 64 possible codons?

There are 64 codons but only 20 amino acids because multiple codons can code for the same amino acid, a feature called redundancy in the genetic code.