SICKLE CELL

Detection of Red Blood Sickle - Cell using DNA Electrophoresis



Detection of Red Blood Sickle - Cell using DNA Electrophoresis

Introduction

Four polypeptides combine to formulate Red blood cells, two chains of alpha and two chains of beta, alpha chains are named as alpha 1 and alpha 2. Beta - globin chain comprises of amino acids and one change in beta - globin series leads to a shape of sickle in the red blood cells. People with African ancestral background are often found with traits of sickle - cell. The red blood cells which have sickle - cell anemia fail to transfer oxygen from lungs to other organs and parts of the body, similarly, they fail to exchange oxygen with carbon - di - oxide to transfer it back to the lungs for filtration and in cases when the level of oxygen or the concentration of oxygen is low in bloods, the outcome is a block in the narrow blood capillaries.

Methods and Materials

The method will utilize six different DNA samples as the material which are named as sample A, B, C, D, E, and F, all belong to different red blood cells. One DNA marker has been provided it is named H. A buffer solution measuring 500 cubic centimeters has been provided which comprises of 10x Tris - borate - EDTA (TBE) solution of stock along with distilled water. The buffer solution has been made in a measuring cylinder of volume 500 cubic centimeter in the ratio of 1 is to 20 for 10x TBE solution to distilled water. The cylinder measurements are that 10x TBE was 25 cubic centimeter and distilled water measured 475 cubic centimeter. This preparation of solution helped in obtaining 0.5 x TBE solutions. A Para film was used to cover the top of the measuring cylinder as it prevented the contact between the hand and the solution when the cylinder was inverted twice or thrice for better mixing of the solution. Agarose gel of 0.8% (w/v) was prepared in a 150 cubic centimeter conical flask where agarose powder weighing 0.24 grams was mixed with 30 cubic centimeter of the diluted buffer solution.

To ensure proper mixing of the solution and powder the conical flask was covered with a loose cotton plug at the top. Heating of the mixture in the conical flask was conducted in a microwave oven on medium settings for dissolving the powder of agarose by boiling the solution, in order to minimize the effect of superheating the flask is swirled. When the entire solid particles of agarose powder mix completely in the solution the solution is left to cool at a temperature of 55 degree Celsius, this is possible as at this temperature human hand can hold the flask for few seconds. Red gel weighing 30 grams is added to the flask mixture because of its characteristics and it will ensure thorough mixing of the solution. The characteristics of red gel are that it is a fluorescent nucleic acid binding dye which by ...