Chapter 19, Problem 13
Describe how the team from the J. Craig Venter Institute created a synthetic genome. How did the team demonstrate that the genome converted the recipient strain of bacteria into a different strain?
Video transcript
If you performed a PCR experiment starting with only one copy of double-stranded DNA, approximately how many DNA molecules would be present in the reaction tube after 15 cycles of amplification?
In a control experiment, a plasmid containing a HindIII recognition sequence within a kanamycin resistance gene is cut with HindIII, re-ligated, and used to transform E. coli K12 cells. Kanamycin-resistant colonies are selected, and plasmid DNA from these colonies is subjected to electrophoresis. Most of the colonies contain plasmids that produce single bands that migrate at the same rate as the original intact plasmid. A few colonies, however, produce two bands, one of original size and one that migrates much less far down the gel. Diagram the origin of this slow band as a product of ligation.
What advantages do cDNA libraries provide over genomic DNA libraries? Describe cloning applications where the use of a genomic library is necessary to provide information that a cDNA library cannot.
You have recovered a cloned DNA segment from a vector and determine that the insert is 1300 bp in length. To characterize this cloned segment, you isolate the insert and decide to construct a restriction map. Using enzyme I and enzyme II, followed by gel electrophoresis, you determine the number and size of the fragments produced by enzymes I and II alone and in combination, as recorded in the following table. Construct a restriction map from these data, showing the positions of the restriction-enzyme cutting sites relative to one another and the distance between them in units of base pairs.
Enzyme Restriction Fragment Sizes (bp)
I 350, 950
II 200, 1100
I and II 150, 200, 950
To create a cDNA library, cDNA can be inserted into vectors and cloned. In the analysis of cDNA clones, it is often difficult to find clones that are full length—that is, many clones are shorter than the mature mRNA molecules from which they are derived. Why is this so?