Difference between revisions of "Project:Blood typing"
Mycoplasma (talk | contribs) |
Mycoplasma (talk | contribs) |
||
| Line 31: | Line 31: | ||
|} | |} | ||
| − | In the UK the distribution of | + | In the UK the distribution of A, B, AB and O is 42%, 10%, 4% and 44%. |
== Process overview == | == Process overview == | ||
| − | 1) Obtain two | + | 1) Obtain two sequences of DNA through PCR, the first containing the deletion site at 258, and the second containing the SNP site at 700 |
| − | 2) Use restriction enzyme KPNI on the first fragment to cut the O alleles only | + | 2) Use restriction enzyme KPNI on the first fragment to cut the O alleles only, and use restriction enzyme ALUI on the second fragment to cut the B alleles only. |
| − | 3) | + | 3) Do gel electrophoresis on the resulting fragments, hopefully resulting in distinguishable bands to show the 6 different alleles. We should then be able to determine an individual's blood type. |
| − | == Process reality | + | == Process reality == |
| + | |||
| + | Much of the necessary equipment we already have from the sex typing experiments. Of the new things, we need the restriction enzymes, and possible a new gel - see below. | ||
All fragments in the papers are between 80 and 200 bp long. For this we would need a polyacrylamide gel (which we have decided against due to difficulty with handling) or a high quality agarose concentrated at 3%, which is a bit expensive, but not impossible. So we are currently looking into the possibility of doing the test using larger fragments. Our agarose at the moment is suitable for fragments > 500bp. | All fragments in the papers are between 80 and 200 bp long. For this we would need a polyacrylamide gel (which we have decided against due to difficulty with handling) or a high quality agarose concentrated at 3%, which is a bit expensive, but not impossible. So we are currently looking into the possibility of doing the test using larger fragments. Our agarose at the moment is suitable for fragments > 500bp. | ||
| + | |||
| + | == New equipment == | ||
| + | |||
| + | Restriction enzymes are listed [here] by sigma: | ||
| + | http://www.sigmaaldrich.com/catalog/search?interface=All&term=restriction+enzymes&lang=en®ion=GB&focus=product&N=0+220003048+219853121+219853286&mode=match%20partialmax | ||
== Sources == | == Sources == | ||
Revision as of 11:55, 15 June 2012
How genes code for blood group
Blood group is determined by the combination of A and B antigens in your red blood cells. A and B individuals have only their respective corresponding antigens, AB individuals have both, and O individuals have none.
The production of these antigens is determined by the "histo-blood group ABO system transferase" gene (1062 base pairs), which is part of the ABO gene locus. This gene codes for the expression of a glycosyltransferase enzyme which by acting on another antigen(H), produces A or B antigens. The A and B allelic forms of the gene code for different forms of glycosyltransferase which affect the H antigen in different ways. O alleles code for another protein that doesn't affect the H antigen, meaning no A or B antigens are expressed. O alleles have a deletion of G at 258, while B alleles have a single nucleotide polymorphism (SNP) from G to A at position 700.
Each person has two of these alleles, one from each parent. A and B are dominant, O is recessive, so the possible combinations are:
| Alleles | Blood group |
|---|---|
| AA | A |
| AO | A |
| BB | B |
| BO | B |
| AB | AB |
| OO | O |
In the UK the distribution of A, B, AB and O is 42%, 10%, 4% and 44%.
Process overview
1) Obtain two sequences of DNA through PCR, the first containing the deletion site at 258, and the second containing the SNP site at 700
2) Use restriction enzyme KPNI on the first fragment to cut the O alleles only, and use restriction enzyme ALUI on the second fragment to cut the B alleles only.
3) Do gel electrophoresis on the resulting fragments, hopefully resulting in distinguishable bands to show the 6 different alleles. We should then be able to determine an individual's blood type.
Process reality
Much of the necessary equipment we already have from the sex typing experiments. Of the new things, we need the restriction enzymes, and possible a new gel - see below.
All fragments in the papers are between 80 and 200 bp long. For this we would need a polyacrylamide gel (which we have decided against due to difficulty with handling) or a high quality agarose concentrated at 3%, which is a bit expensive, but not impossible. So we are currently looking into the possibility of doing the test using larger fragments. Our agarose at the moment is suitable for fragments > 500bp.
New equipment
Restriction enzymes are listed [here] by sigma: http://www.sigmaaldrich.com/catalog/search?interface=All&term=restriction+enzymes&lang=en®ion=GB&focus=product&N=0+220003048+219853121+219853286&mode=match%20partialmax
Sources
Most of the procedure came from this paper:
http://library-resources.cqu.edu.au/JFS/PDF/vol_41/iss_6/JFS416961027.pdf
With some more here: