Lab 03B: HHMI Bacterial Identification Virtual
Lab questions with verified answers
after the final step of PCR purification, what should be in the tube
Ans✓✓✓many copies of 16s rDNA, each about 15000 base pairs (bp)
long
after you spin the inverted column at 3000 rpm for 2 minutes, where is
the PCR samples Ans✓✓✓in the collection tube
describe the DNA in each strip tube Ans✓✓✓All DNA pieces in each
tube start with the same primer but end with different nucleotide tagged
with a fluorescent marker
Describe the process of extracting bacterial DNA (sample preparation)
Ans✓✓✓1. dissolve the cell wall with digestive buffer
2. heat sample in a water bath 100°C to denature proteolytic enzymes
from digestive buffer
3. spin sample in centrifuge
4. transfer supernatant (the liquid) to PCR tube
describe the steps of PCR Ans✓✓✓1. add PCR Master Mix solution to
sample DNA, positive control, and negative control
2. add positive and negative controls to their tubes
3.load tubes into thermocycler (PCR machine), run and then remove
, 4. insert microconcentrator column of appropriate size into collection
tubes
5. add 400µL of buffer to columns
6. add entire PCR content (~100µL) to column
7. put positive and negative control tubes on ice
8. Spin column at 3,000 rpm in a fixed-angle centrifuge for 15min
9. Remove column and discard collection tube
10. invert column, attach to new collection tube, add 50µL of buffer to
inverted column
11. spin inverted column 3,000rpm for 2min
12. discard column
describe what happens in PCR Ans✓✓✓In normal cells, the dsDNA is
unzipped with an enzyme to start the replication process. In PCR,
ssDNA is made by heating a chromosome fragment to 95°C. It is then
cooled.
describe what is happening in one tube containing the primer 651R
during sequencing prep Ans✓✓✓each DNA strand binds the primer at
one end and will have a fluorescence-tagged terminator at the other hand
explain what happens in an automatic sequencer Ans✓✓✓the sequencer
has a thin capillary tube attached at one end to a syringe mechanism that
contains a buggger solution. The tube is filled with the bufferr solution
and the other end inserted into one of the tubes contains the DNA pieces.
An electric current is applied so that the end of the tube in contact with
the DNA has a negative charge and the syringe end a positive charge.
Lab questions with verified answers
after the final step of PCR purification, what should be in the tube
Ans✓✓✓many copies of 16s rDNA, each about 15000 base pairs (bp)
long
after you spin the inverted column at 3000 rpm for 2 minutes, where is
the PCR samples Ans✓✓✓in the collection tube
describe the DNA in each strip tube Ans✓✓✓All DNA pieces in each
tube start with the same primer but end with different nucleotide tagged
with a fluorescent marker
Describe the process of extracting bacterial DNA (sample preparation)
Ans✓✓✓1. dissolve the cell wall with digestive buffer
2. heat sample in a water bath 100°C to denature proteolytic enzymes
from digestive buffer
3. spin sample in centrifuge
4. transfer supernatant (the liquid) to PCR tube
describe the steps of PCR Ans✓✓✓1. add PCR Master Mix solution to
sample DNA, positive control, and negative control
2. add positive and negative controls to their tubes
3.load tubes into thermocycler (PCR machine), run and then remove
, 4. insert microconcentrator column of appropriate size into collection
tubes
5. add 400µL of buffer to columns
6. add entire PCR content (~100µL) to column
7. put positive and negative control tubes on ice
8. Spin column at 3,000 rpm in a fixed-angle centrifuge for 15min
9. Remove column and discard collection tube
10. invert column, attach to new collection tube, add 50µL of buffer to
inverted column
11. spin inverted column 3,000rpm for 2min
12. discard column
describe what happens in PCR Ans✓✓✓In normal cells, the dsDNA is
unzipped with an enzyme to start the replication process. In PCR,
ssDNA is made by heating a chromosome fragment to 95°C. It is then
cooled.
describe what is happening in one tube containing the primer 651R
during sequencing prep Ans✓✓✓each DNA strand binds the primer at
one end and will have a fluorescence-tagged terminator at the other hand
explain what happens in an automatic sequencer Ans✓✓✓the sequencer
has a thin capillary tube attached at one end to a syringe mechanism that
contains a buggger solution. The tube is filled with the bufferr solution
and the other end inserted into one of the tubes contains the DNA pieces.
An electric current is applied so that the end of the tube in contact with
the DNA has a negative charge and the syringe end a positive charge.
