BIO 208 Prelab:PCR of HUMAN ALU ELEMENT S09

1

BIO 208 PreLAB:PCR of HUMAN ALU ELEMENT s09

*Use separate piece of paper. Type or write neatly. Use full sentences.

Recommended reading:

Alu elements and transposons, Chapter 2, page 36 – 37

Polymerase chain reaction, Chapter 8, page 200 – 202 and figure 8.21

Handout from class

1. a. Approximately what % of the genome is composed of Alu elements?

b. The Alu element you will study is 300 base pairs in length. How many copies of Alu elements are in the human genome?

c. Are Alu elements considered to be coding, or non-coding sequences (choose one)?

d. What is the specific name of the Alu element PCR-amplified in lab? On which chromosome is it located?

1. A few Alu elements are dimorphic. What is meant by this, how are dimorphic elements useful in DNA fingerprinting, and why is a monomorphic element not useful for DNA fingerprinting?

1. A chromosome that contains an Alu element is considered to be “+”. The same chromosome without the Alu element is considered to be “-“. Every individual has 2 copies of each chromosome, one inherited from the mother and one from the father. In a population of chimpanzees there is an infant of unknown paternity. The infant has the Alu genotype of +/+ while the mother is +/ - .

Examine the gel results below. Which of the following chimps could be the father (identify one or more individuals)? What is the basis for your identification? Which, if any of the males could not be the father?

Lane 1Lane 2Lane 3Lane 4Lane 5

Possible fathersWatson SamNobel InfantMother

941 base pairs (+)

641 base pairs (-)

1. List 4 components required for DNA synthesis in the PCR (polymerase chain reaction)

1. Describe (briefly) what occurs at each of the following temperatures in the PCR

94oC for 1 minute  60oC for 1 minute -> 72oC for 1 minute. (Note, the textbook indicates that a temperature from 37 oC to 65 oC can be used, the exact temperature has to be determined for each set of primers, the ones for this particular laboratory "work" at 60 oC)

6. a. What organism is taq polymerase isolated from (include genus and species names)?

b. Where (what type of environment) does this organism live?

c. What does the taq enzyme offer to the PCR that other bacterial polymerases don't?

over 

7. A DNA preparation yields 1 nanogram of DNA. How many cycles of the PCR are required to obtain at least 300 ng of DNA, enough to visualize on an agarose gel? Remember, each PCR cycle doubles the amount of DNA.

8. Optional (not required to turn in as homework) – we will cover this question prior to the lab, it is a critical component of the PCR. What is the role of primers in the PCR and how do primers ensure that only a specific section of the chromosome (the Alu element in this case) is amplified?