Dna

Wednesday, April 4th 2006

By: Jacquelyn Logan

Lab Partners:

Jessica Amedick

Josh Fucinski

Jade Gianforte

ABSTRACT: Starting with whole onions and then ending with a relatively pure preparation of DNA by isolating it from living plant tissue. By knowing the physical and biochemical characteristics of the genetic material the isolation of DNA is possible. In the first steps of homogenization the lysis medium containing detergent caused the cell membrane to break down and release the l

INTRODUCTION:

The instructions that tell a living organism how to develop, grow, or what molecules to produce and when are contained in the nucleus of every cell of that particular organism. The cell’s nucleus contains DNA, which stores genetic information. In eukaryotes, such as those in onion cells, DNA is packaged into structures called chromosomes. Each chromosome consists of a single large DNA molecule that contains thousands of different genes depending on the organism.

A DNA molecule is a polymer composed of many repeating subunits called nucleotides. Each nucleotide is made up of a 5-carbon sugar, a phosphate, and a nitrogenous base. How the bases connect together is what gives the nucleotide it’s specific characteristics. The genetic information encoded in the DNA molecule is determined by the sequence of nucleotides in the chain.

Science has been able to increase it’s knowledge of the structure and function of chromosomes and genes, and has been able to utilize that knowledge in biotechnology, thanks to the development of suitable techniques. One important technique has been the extraction and isolation of DNA from living organisms.

Isolation of DNA is done in three basic steps. First, the addition of the detergent and the mashing of the onion, to break down the cell walls and to dissolve the fat molecules that make up the cell wall. This allows the DNA to be released. Heating the mixture precipitates the lipids (fats) and proteins out of solution. The addition of the cold alcohol precipitates the DNA from the solution since DNA is insoluble in alcohol.

When extracting DNA from any organism, the cell walls and cell and nuclear membranes must be broken. Once the structural components of the cells are released, we can use the chemical properties of all the cell contents to separate the DNA from all the other molecules contained within the cells. DNA isolation involves several steps including homogenization and precipitation. Each of the steps requires specific chemicals, techniques, and procedures that must be carefully followed.

Homogenization is the first step in isolation of the onion cells. In this step, we will break up the onion tissues to separate and open the cell. The cell walls and nuclear membranes will get fragmented, and the molecules contained within the cells (including the DNA) are released into the medium. By briefly heating the tissue before homogenization will help inactivate DNAases (enzymes that degrade DNA) that are present in our mixture.

Before DNA can be released from the nuclei the cell walls, plasma membranes, and nuclear material must first be broken down. This can be accomplished by homogenizing the onion tissues in a blender. The lysis medium, which contains detergent, causes the cell membrane to break down and emulsifies the lipids and proteins of the cell by disrupting the polar interactions that hold the cell membrane together.

MATERIALS & METHODS:

Refer to Lab Nine: Isolation of DNA from Onion, (Titus 2006) on Tuesday, April 4th.

DATA & RESULTS:

In the first steps in the homogenization released the DNA from the nuclei of the onion tissues, the cell walls, and plasma membranes. The foam that formed during the blending portion of the homogenization step was the proteins precipitation out of the solution, causing the DNA to become separated from the chromosomal proteins.

After the blending