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ACID HYDROLYSIS FROM THE ISOLATED PLANT DNA

Lanzona, Marionne., Laquindanum, Maan., Libed, Karessa C., Lim, Billy., Madrigal, Richelle., Naui, Mikhaela Group 5 2F Pharmacy General Biochemistry Laboratory

ABSTRACT
Cells in our world come in 2 basic types prokaryotic and eukaryotic. All plant and animal cells belong to the type eukaryotic. The most distinctive feature of the eukaryotes is that they have a cell nucleus. On the other hand, some single celled organisms such as bacteria have no cell nucleus and the DNA molecule is free inside the cell. This type is called the prokaryotic. DNA is unique to each individual like fingerprints. Thus DNA can be mapped to reveal the genetic makeup of an organism. The technique of DNA mapping or fingerprinting was discovered by Alec J. Jeffreys in 1984. DNA fingerprinting in plants is used for protection of the ecosystem, identifying marker traits, identification of gene diversity and variations and mutations.

INTRODUCTION
Deoxyribonucleic acid or DNA, is a nucleic acid that contains the generic instructions used in the development and functioning of all known living organisms. The main role of DNA molecules is the long-term storage of information. It contains the instructions needed to construct other components of cells. DNA segments that carry this genetic information are called genes. DNA consists of two long polymers called nucleotides, with backbones made of sugar and phosphate groups joined by ester bonds. These two strands run in opposite directions to each other and are therefore anti-parallel. DNA is stable to bases (alkaline hydrolysis). This is one of the features which distinguish DNA from RNA (RNA is not stable under alkaline conditions). Strong acids at a high temperature, are capable of breaking the DNA molecule into its components. These conditions break both of the phosphate ester bonds and also the N-glycosidic bond between the deoxyribose and the purines and pyrimidine bases. DNA hydrolysis is the breaking of DNA through the addition of water. It can be done in several fashions; enzymatic (exonucleases)or chemicals (acid). The main difference between plant and animal DNA is in the organization of genes and the size of the total genome or how many base pairs of DNA the organism has. Plants tend to have larger genomes than animals. Hydrolysis is for the next test which is the chemical characterizations. The DNA must be thoroughly isolated and adjusted to the right pH beforehand.

EXPERIMENTAL A.COMPOUNDS TESTED


The following items and/or reagents were used in the experiment: Red onions, 1M HCl, 1M KOH and Glacial acetic acid.

B.PROCEDURE
After the isolation of the plant DNA, a small amount of the DNA sample was added to 1 mL of 1M HCl. The mixture was covered with parafilm and then heated to 100 for 60 minutes with

occasional agitation. After heating, it was cooled with running water. The hydrolysate was adjusted or neutralized to pH 4 then centrifuged and decantated.

RESULTS AND DISCUSSIONS


After the isolation of the plant DNA, the suspension was turbid. A small amount was added to a mixture, while the mixture was being heated, we observed the recoiling of the DNA or there were strands appearing. After heating, it was neutralized or adjusted to pH 4. To determine if its the correct pH, we used a colored pH paper (as seen on figure 2). After all of that, it was centrifuged and decantated.

From the internet: http://www.massey.ac.nz/~wwbioch/DNA/hydDN A/framset.htm (February 8, 2011) http://www.tutorvista.com/biology/plant-dna (February 8, 2011)

Figure 1. Isolated plant DNA

Figure 2. pH paper (pH 4)

Figure 3. After the acid hydrolysis

REFERENCES
From books: Torres, Peter C., et. Al. (2010) Laboratory Manual in General Biochemistry. C & E publishing inc. Boyer, Rodney. (2006) Concepts in Biochemistry 3rd ed. C & E publishing inc.