We are trying to detect the prevalent mutation in case of sickle cell anemia in a specific population. We are confused about which gene we should consider as we were unable to find much literature on it. Could someone help us with this problem? What are the restriction enzymes that can be used? Where can we get them? Thank you
Dear Ashish Bhandari,
This may help you:
Mutations in the globin genes that alter the protein composition but not necessarily the amount of expression are referred to as qualitative mutations. Of the mutations leading to qualitative alterations in hemoglobin, the missense mutation in the β-globin gene that causes sickle cell anemia is the most common. The mutation causing sickle cell anemia is a single nucleotide substitution (A to T) in the codon for amino acid 6. The change converts a glutamic acid codon (GAG) to a valine codon (GTG). The form of hemoglobin in persons with sickle cell anemia is referred to as HbS. The nomenclature for normal adult hemoglobin protein is HbA.
The underlying problem in sickle cell anemia is that the valine for glutamic acid substitution results in hemoglobin tetramers that aggregate into arrays upon deoxygenation in the tissues. This aggregation leads to deformation of the red blood cell into a sickle-like shape making it relatively inflexible and unable to traverse the capillary beds. This structural alteration in the red blood cell can easily be seen under light microscopy and is the source of the name of this disease. Repeated cycles of oxygenation and deoxygenation lead to irreversible sickling.
http://themedicalbiochemistrypage.org/sicklecellanemia.php
Your question is rather puzzling! There is only one mutation giving rise to the sickle hemoglobin (HbS) : beta 6(A3) Glu>Val or HBB:c.20A>T. The mutation abolishes a site for the restriction enzymes Dde I , Mst II or Bsu 361 (an isoschizomer of Mst II).
Thank you very much for your response. I am also sorry for the confusion. I was clear that there was a single mutation involved in sickle cell anemia but I could not find the gene to look for in NCBI database. My question was to address the issue of us not being able to locate the gene in the NCBI database as there are many entries in NCBI database referring to the gene. Could you help me with that.
Is this what you are looking for?
Medical Intelligence
A Sensitive New Prenatal Test for Sickle-Cell Anemia
Judy C. Chang, M.S., and Yuet Wai Kan, M.B., D.Sc.
N Engl J Med 1982; 307:30-32July 1, 1982DOI: 10.1056/NEJM198207013070105
The gene is the Beta-globin gene located on chromosome 11. The answer of Jacques Elion is complete regarding the restriction enzymes.
Yes indeed, there are several entries for the beta-globin gene (HBB) in several databases
One you may look at is Ensembl:
HBB ENSG00000244734
http://www.ensembl.org/Homo_sapiens/Gene/Sequence?g=ENSG00000244734;r=11:5225464-5229395
Are you planning to do prevalence studies in specific population for sickle cell anemia ? or you wants to do pre-natal diagnosis ?
As this PCR based test is molecular diagnostic test, it is costly for developing countries including India and countries where Indo Arab haplotype is very common.
Why dont you go for simple protein based identification by established method such as solubility test followed by Hb electrophoresis on cellulose acetate membrane. This combined test is the low cost golden standard for the diagnosis of sickle cell anemia.
Whereas, PCR based test is costly and only routinely used for pre natal diagnosis.
Hope this will give you second opinion on which test to be used for what purpose.
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