Chapter 20: DNA Technology and Genomics
1. What is recombinant DNA?

2. An example of recombinant DNA is demonstrated by the modern production of insulin:
Insulin is a simple protein normally produced by the pancreas. In people with diabetes, the pancreas is damaged and cannot produce insulin. Since insulin is vital to the body's processing of glucose, this is a serious problem. Many diabetics, therefore, must inject insulin into their bodies daily. Prior to the 1980s, insulin for diabetics came from pigs and was very expensive.

To create insulin inexpensively, the gene that produces human insulin was added to the genes in a normal E. coli bacteria. Once the gene was in place, the normal bacteria cellular machinery produced it just like any other protein. Large quantities of the modified bacteria are cultured and then the insulin is extracted, purified and used very inexpensively. (for more information see http://science.howstuffworks.com/cell13.htm and http://www.howstuffworks.com/diabetes.htm)
Question: If you were a scientist that wanted to make a recombinant plasmid (that would eventually be taken up by bacteria through transformation) describe the steps using these terms in this order... cloning vector, gene of interest (insulin gene), restriction site, restriction enzymes, sticky ends, complementary sticky ends, ligase, transformation, recombinant bacteria containing the human insulin gene.

3. But it’s not really as simple as question #2 describes... to get a eukaryotic gene like insulin to actually function within a prokaryotic organism the following must also be done to that eukaryotic gene:
a. cDNA must be used. What is cDNA --AND-- why must we use cDNA in the bacterial
transformation process?
b. Also, why do we need to insert a prokaryotic promoter right before the eukaryotic gene of
interest?

4. Geneticists need ways to be able to determine if a bacteria cell has actually received the gene of interest. Describe how these 3 processes could be used to do that:
a. What process was used in fig. 20.4
b. nucleic acid probe hybridization
c. DNA gel electrophoresis

5. a. If, as a crime scene detective, you were able to collect some very small samples of skin from under a suspect’s fingernails, why would you want to use a PCR reaction on your samples?
b. How does PCR work?

6. Gel electrophoresis:
a. Why does DNA electrophoresis require restriction enzymes?
b. Why does DNA move toward the positive electrode in a gel box?
c. Why are the larger DNA fragments closer to the negative electrode than the smaller fragments are?

7. Describe or draw what a finished gel would look like if it were cut at the following sites:

__/______/____/______/______
9 71 14 196 50

8. Southern blotting uses nucleic acid probe hybridization technology similar to fig. 20.5.
a. Why are the results at step 2 of fig.20.10 different than the results at steps 4&5 of the same figure?
b. What do the results (step 5) for samples I, II, and III tell you about the three individuals tested?

9. Micro-array assay (a definition for assay is: a technique for surveying concentrations or compositions of a substance... in this case tissues are surveyed for the proteins they produce)
a. Describe hoe DNA microarray assays are performed.
b. If you compared pancreas tissue and muscle tissue with this technique what obvious differences in gene expression (protein production) would you expect to see?

10. Using p 402-408 list at least 10 ways in which DNA technology does or could one day eventually affect your own life.

Chapter 21 The Genetic Basis of Development

1. a. Why are fruit flies an excellent model organism for genetic studies?
b. Why might mice or zebra fish be more suitable than flies for some studies related to humans?

2. a. Define morphogenesis
b. Define cell differentiation

c. give an example of each of the terms above using figure 21.4

3. Thoroughly describe the differences between human adult stem cells and embryonic stem cells?

4. Using the textual information and figure 21.5 on p.415:
a. explain totipotency in plants.
b. explain what is meant by dedifferentiation.

5. In the cytoplasm of an egg cell there are maternal substances such as mRNA, proteins, hormones etc. These substances/chemicals influence very early stages of embryonic development.
a. These substances are called ______.
The substances are unevenly distributed and after fertilization mitosis occurs which begins to form new cells, each with differing concentrations and types of chemicals. These substances can serve as early transcription factors and controlling elements, which cause different cells to express different genes and eventually become different tissues in the developing embryo.
b. What does induction have to do with the above processes?

6. Define:

a. maternal effects genes

b. gap genes

c. pair-rule genes

d. segment polarity genes
7. a. What are homeotic genes?
b. What does the book mean by the term “master regulatory switches”?

8. a. What is apoptosis?
b. How is apoptosis involved in the formation of separate digits in all mammals and land birds?

c. What other reasons are there for apoptosis to occur in humans?

9. Homeotic genes (of the homeobox) are high conserved throughout the animal kingdom.
a. What does that mean?
b. Why might these homeotic genes be more conserved between species than most other genes of the genome?