You have generated three transgenic lines of maize that are resistant to the European corn borer, a significant pest in many regions of the world. The transgenic lines (T₁ in the accompanying table) were created using Agrobacterium-mediated transformation with a T-DNA having two genes, the first being a gene conferring resistance to the corn borer and the second being a gene conferring resistance to a herbicide that you used as a selectable marker to obtain your transgenic plants. You crossed each of the lines to a wild-type maize plant and also generated a T₂ population by self-fertilization of the T₁ plant. The following segregation results were observed (herbicide resistant : herbicide sensitive):

Explain these segregation ratios.

In Drosophila, loss-of-function Ultrabithorax mutations result in the posterior thoracic segments differentiating into body parts with an identity normally found in the anterior thoracic segments. When the Ultrabithorax gene was cloned, it was shown to encode a transcription factor and to be expressed only in the posterior region of the thorax. Thus, Ultrabithorax acts to specify the identity of the posterior thoracic segments. Similar genes were soon discovered in other animals, including mice and humans. You have found that mice possess two closely related genes, Hoxa7 and Hoxb4, which are orthologs of Ultrabithorax. You wish to know whether the two mouse genes act to specify the identity of body segments in mice.

How will you determine where and when the mouse genes are expressed?

Bacterial Pseudomonas species often possess plasmids encoding genes involved in the catabolism of organic compounds. You have discovered a strain that can metabolize crude oil and wish to identify the gene(s) responsible. Outline an experimental protocol to find the gene or genes required for crude oil metabolism.

Two complaints about some transgenic plants presently in commercial use are that (1) the Bt toxin gene is constitutively expressed in them, leading to fears that selection pressures will cause insects to evolve resistance to the toxin, and (2) a selectable marker gene—for example, conferring kanamycin resistance—remains in the plant, leading to concerns about increased antibiotic resistance in organisms in the wild. How would you generate transgenic plants that produce Bt only in response to being fed upon by insects and without the selectable marker?

You have identified five genes in S. cerevisiae that are induced when the yeast are grown in a high-salt (NaCl) medium. To study the potential roles of these genes in acclimation to growth in high-salt conditions, you wish to examine the phenotypes of loss- and gain-of-function alleles of each. How would your answer differ if you were working with tomato plants instead of yeast?

You have isolated another cDNA clone of the CRABS CLAW gene from a cDNA library. The cDNA was directionally cloned using the EcoRI and XhoI sites. You sequence the recombinant plasmid using primers complementary to the T7 and T3 promoter sites flanking the MCS. The first 30 to 60 bases of sequence are usually discarded since they tend to contain errors.

Can you identify the start of the coding region in the end of the gene? What does the sequence preceding the start codon represent?

You have identified five genes in S. cerevisiae that are induced when the yeast are grown in a high-salt (NaCl) medium. To study the potential roles of these genes in acclimation to growth in high-salt conditions, you wish to examine the phenotypes of loss- and gain-of-function alleles of each. How will you do this?