Exam 17: Applications of Recombinant Dna Technology and Reverse Genetics

Snuppy, a male Afghan hound, was created by transferring the nucleus from an ear cell of the male Afghan, Tai, into an enucleated egg from a donor. The embryo was transplanted into a surrogate mother to develop to term. Which of the following animals would be a genetic match for Snuppy if nuclear DNA were tested?

Current sequencing technologies have major advantages over the dideoxy method because they produce more sequence in less time and at less cost. Which of the following would provide further advantage?

If genes are introduced into a yeast plasmid to serve as a shuttle vector, the plasmid has the ability to __________.

Some restriction enzymes cut at specific and unambiguous sequences (e.g., NotI at GCGGCCGC). As such, the distance between two such sites in a large random DNA sequence is 4⁸, assuming a 50% G+C DNA content. However, the sequence recognized by XhoII is said to be ambiguous (Pu-GATC-Py, where Pu = any purine and Py any pyrimidine). What, therefore, is the expected distance between two XhoII sites in a large random sequence?

For a transgenic animal line to be produced from a chimera, the transgene must be carried by what?

Suppose you have engineered a plasmid vector in order to produce a functional animal protein in E. coli. But the product you collect, while it has the correct amino acid sequence, is in fact nonfunctional. What might you try that would have a greater chance of success?

If a gene is replaced with a heterologous sequence such that the gene is then nonfunctional, what is the result usually called?

In using Agrobacterium tumefaciens to transfer genes into plants, what is transferred from bacterium to plant?

In creating a recombinant DNA molecule, what are two methods that prevent re-ligation of the cloning vector that contains no insert DNA?

In the 1990s, gene therapy was seen as a probable cure for many genetic diseases. List three or four reasons that it has so far been of limited success.

Whether libraries are cDNA or genomic, whether amplification is based on PCR or cloning, and whether screening is done with northern or Southern blotting or probes, are all decisions based on which of the following?

A hypothetical restriction sequence is 5′-TCTAGA-3′. The palindromic sequence is 3′-________-5′.

In a cDNA library, not all expressed genes will be represented. On what does this depend?

In producing a recombinant plasmid to be used to clone a given donor insert, it is possible to cut both the donor and plasmid with the same restriction enzyme, resulting in complementary sticky ends. Assuming plenty of plasmid DNA is available, why is further selection necessary before the introduction of the plasmid into a cellular system?

In grape cultivation, growers often find that heterozygous forms are more desirable. To propagate the heterozygotes without having to interbreed in each generation, what method can they use?

Your student is trying to create a "Gene X" knockout mouse by introducing a recombinant construct into embryonic stem (ES)cells, selecting transformants, and introducing the transformed cells into mouse blastocysts. He designed his construct as follows: Neo^R----region of homology to Gene X---HSV tk--- region of homology to Gene X where Neo^R is the neomycin-resistance gene and HSV tk is the Herpes Simplex virus thymidine kinase gene. He transforms the ES cells and exposes them to G418 and ganciclovir. Explain to your student he failed to recover transformed ES cells.

In the formation of transgenic animals, DNA is usually inserted into eggs or embryos. If some cells of the embryo do not receive the introduced DNA, the result is an organism with cells of different genotypes; the organism is known as what?

You have a DNA sequence of 1600 bp. To characterize it, you decide to make a restriction map. Cutting with the first enzyme, you show by electrophoresis that the fragments produced are 400 and 1200 bp, respectively. After using the second enzyme, you recover 700-bp and 900-bp fragments. After treatment with both enzymes, fragments were 300, 400, and 900 bp. Which is the correct map?

If recombination results in random integration of a DNA sequence so that it produces a non-homologous region, the recombination is ________.

Describe two or more ethical issues arising from sequencing the genomes of individuals.