Follow the prompts to put on gloves and pick up and move items as needed to collect a bacterial colony. Why do many pathogenic bacterial samples grow poorly on solid culture medium?

HHMI Biointeractive Worksheet
Name:
Follow the link to open the HHMI Biointeractive Unknown Bacterial Identification Tab.
https://www.biointeractive.org/classroom-resources/bacterial-identification-virtual-lab
Click on the Launch Interactive Button:

⦁ What are the Basic Steps to identify different types of bacterial based on their DNA sequence?

⦁ What is 16s rDNA and how is it used to identify organisms?

⦁ Click to enter the Lab and then click on the Sample Tab to view where the samples you are testing were collected from.

Where were the 6 samples collected from?

⦁ Follow the prompts to put on gloves and pick up and move items as needed to collect a bacterial colony. Why do many pathogenic bacterial samples grow poorly on solid culture medium?

⦁ The first step is to extract the bacterial DNA by dissolving the cell wall with proteolytic enzymes. Why do you need to get rid of the proteolytic enzymes used to degrade the cell wall prior to adding the other enzymes?

⦁ After you denature the enzymes and centrifuge the sample, where is the DNA located? Is it located in the pellet or the supernatant?

⦁ Once you have the DNA then you must complete a PCR Amplification. What is in the master mix you added to your DNA sample?

⦁ What is happening during each cycle of the of the PCR Run?

⦁ The 3rd part is the purification of the PCR Product. What is found in the tube at this point?

⦁ How many base pairs will the amplified DNA contain?

⦁ Part 4 is Preparing the Sequencing Samples. What is the “sequencing brew” that is found in the Green and blue tubes?

⦁ Part 5 is the DNA sequencing. How are the different sized fragments separated to determine the sequences?

⦁ The sequencer reads the color on of fluorescences on the end of each fragment. What color is associated with the A? the T? the G? the C?

⦁ Part 6 – DNA Sequence Analysis. What is BLAST?

⦁ Click the link to get the Sequence Data and then BLAST the sequence using the NCBI BLAST tool. What is the identity of the organism in Sample A?

⦁ Click on the Sample Tab and go to Sample B. Collect the sequence data and BLAST the sequence to identify the organism in Sample B.

⦁ Repeat the samples to identify Sample C.

⦁ Identify Sample D

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