Dissociation%20Polynucleotides%20_Fall%202009_

Dissociation%20Polynucleotides%20_Fall%202009_ - BME 100L...

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1 BME 100L. Modeling Cellular and Molecular Systems Fall 2009 A Thermodynamic Lab on Dissociation of Double Stranded Polynucleotides Lab report is due 5pm (11/23 for Group A) and (12/02 for Group B) Objective: Inosinic acid (I) is a derivative of adenosine and can bind to cytidine (C), adenosine (A), and uridine (U). In this lab, we will determine the dissociation of double-stranded chains of polyinosinic acid-polycytidinic acid (polyI-polyC) as a function of temperature. “Dissociation” means breaking hydrogen bonds between complementary double-stranded nucleotide chains, resulting in two single-stranded chains. The structure of polyI-polyC is shown below. Note the locations of hydrogen bonds. O O P OH OH O OH OH N N N NH Inosinic acid Cytidinic acid O O P H N 2 Hydrogen bonds The dissociation can be measured by taking advantage of the hypochromicity of the polynucleotides (i.e. increased light absorption as double-stranded chains become single- stranded ones when the solution is heated). Background: Nucleotides in each polynucleotide strand are held together by covalent phosphodiester bonds that link the pentoses of adjacent nucleotide via the phosphate group. This is often called the “sugar-phosphate backbone” of the polynucleotides. The polynucleotides have directions: the 3’ end (i.e., the hydroxyl terminus on the sugar group) and the 5’ end (i.e., the phosphate terminus). DNA primary structure is formed by the backbone and the following nitrogenous bases: Guanine (G), Adenine (A), Cytosine (C), and Thymine (T).
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2 Similarly, RNA primary structure is also formed by the backbone and nitrogenous bases. However, Uracil is used instead of Thymine in RNA. (Figure from http://ghr.nlm.nih.gov) The most thermodynamically stable conformation of polynucleotides is in double- stranded form, where complementary polynucleotides are paired by hydrogen bonds. “Complementary” refers to the nitrogenous bases that form hydrogen bonds with each other. In Thymine Guanine Adenine Cytosine Uracil (Figures from the Nucleic Acid Database http://ndbserver.rutgers.edu)
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3 DNA, A and T are complementary (form two hydrogen bonds between them), and G and C are complementary (form three hydrogen bonds between them). This is often written as A::T and G:::C. The hydrogen bonding is called “base pairing.” As a result, the famous double helix structure is formed (see the figure above). Although RNA is often a single-stranded molecule, it is more thermodynamically stable
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This note was uploaded on 01/17/2010 for the course BME 100 taught by Professor Yuan during the Fall '07 term at Duke.

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Dissociation%20Polynucleotides%20_Fall%202009_ - BME 100L...

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