CRIM 101 Lecture 34: CRIM LEC34

What is shown below for each is only the beginning of each part. Please finish each part entirely

Briefly describe the purpose or function of the DNA sequences in the pETDuet-1 Vector listed below.

1. Ampicillin resistance gene

2. ORI

3. lacI gene

4. T7 promoter

5. lac operator

6. rbs

7. His-Tag

8. MCS

9. Start codon

10. pET Upstream Primer

1.In the T7/lac expression system used for ADH expression, which of the following is true?

The promoter for the ADH gene is specific for E. coli RNA polymerase.

The promoter for the ADH gene contains the lac operator sequence.

The pLacI plasmid encodes the T7 RNA polymerase gene.

All of the above

2.The ADH gene used in this lab contains

A his-tag at the N-terminus

An HSV tag near the N-terminus

A strep tag at the C-terminus

None of the above

3.During ADH expression in Tuner cells, which of the following occurs?

Binding of IPTG to the Lac repressor protein facilitates expression of the T7 RNA polymerase.

IPTG represses the expression of the Lac repressor protein.

The presence of glucose in the medium results in CAP-binding protein recruiting E. coli RNA polymerase to the T7 RNA polymerase gene.

All of the above

4. How would you interpret your NovaBlue transformation results if you saw no colonies on any of your plates (controls and experimental)?

You used the wrong cells for the transformation.

You used the wrong medium for the transformation.

You heat shocked the cells for too long.

Any of the above could explain the results.

5.In SDS-PAGE, which of the following is true?

The proteins are only negatively charged in the resolving gel.

The stacking gel has a higher pH than that of the resolving gel.

The concentration of acrylamide is higher in the stacking gel relative to the resolving gel.

none of the above