1)How can you indicate that the bacteria have taken up the plasmid?

2)What is the purpose of the LB/amp(-) plate? Should there be bacterial growth on the plate?

3) Why would one compare plates LB/amp (-) and LB/amp (+)?

1) To indicate that the bacteria have taken up the plasmid, you can use a method called plasmid DNA sequencing or colony PCR. These methods involve isolating DNA from the bacterial colonies and analyzing it for the presence of the target sequence or amplicon.

For plasmid DNA sequencing, you would need to extract DNA from the bacterial colonies and send it for sequencing. The sequencing results can then be analyzed to determine if the target sequence of interest is present in the extracted DNA.

Alternatively, for colony PCR, you would need to select a few bacterial colonies and directly amplify the target sequence using PCR. The PCR products can then be analyzed on an agarose gel to check if the expected amplicon size is present.

2) The LB/amp(-) plate is a selective medium that is used to select for bacteria that have taken up the plasmid containing the ampicillin resistance gene. Ampicillin is an antibiotic that inhibits the growth of bacteria lacking the ampicillin resistance gene. Therefore, the purpose of the LB/amp(-) plate is to identify and select bacterial colonies that have successfully incorporated the plasmid.

Ideally, there should not be any bacterial growth on the LB/amp(-) plate, except for the bacteria that have taken up the plasmid and acquired resistance to ampicillin. This indicates that the antibiotic selection has worked, and only transformed bacteria can grow on the plate.

3) Comparing the LB/amp(-) and LB/amp(+) plates allows you to determine the efficacy of plasmid transformation.

The LB/amp(-) plate serves as a negative control. It should only have the transformed bacteria growing on it because they possess the ampicillin resistance gene carried by the plasmid. Any growth on the LB/amp(-) plate indicates successful transformation.

On the other hand, the LB/amp(+) plate is a positive control. It contains the same antibiotic, ampicillin, but without the plasmid. This plate should not have any bacterial growth. If there is any growth present on the LB/amp(+) plate, it suggests that there may be contamination, such as spontaneous mutations in the bacteria or issues with the antibiotic itself.

By comparing these two plates, you can confirm the specificity of the ampicillin resistance gene and evaluate the success of your transformation experiment.