Bio 362 Human Genetics, Spring 2008
Sickle cell anemia study sheet
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Web reading:
Why would you consider this a reputable web site?
What is the molecular structure of hemoglobin?
What hemoglobin variants exist in a normal person, and at what life stages do they appear?
What genes encode components of hemoglobin?
What mutant varieties of hemoglobin exist?
What happens to HbS during an oxygenation/deoxygenation cycle? How does damage occur?
How does hydroxyurea help sickle cell patients? How is this mechanism related to differing disease severity in different areas?
What explanations have been proposed for how sickle cell heterozygosity protects against malaria?

Ingram, 2004 :
This paper is a commentary. The purpose is not to relate or summarize new results, but instead to convey a personal perspective and/or new hypothesis.
By what methods did Pauling, initially, and then Ingram and colleagues deduce the structural difference between HbA and HbS?
Why was it important to examine HbS from five different patients?
How did Ingram's results contribute to the emerging knowledge of how DNA sequences specify protein sequences?
What's the difference between sickle cell anemia and thalassemias? (see also background readings)
How might you explain the allele frequencies of HbS and HbC in Suriname and Curacao in 1965? (see bottom of right column, page 4)
Explain the theme of serendipity.

Walters, 2005:
This is a review paper. Results of many primary research papers are summarized.
What type of cell replacement therapy currently works for sickle cell anemia?
What usually causes the death of an allograft (from another person) cell transplant recipient?
What other limitations restrict the applicability of cell transplanation via allograft?
What is chimerism? What are the findings regarding chimerism in allograft transplant recipients?
What are the implications of the chimerism findings for the possible success of gene therapy techniques?
How are cells harvested for gene therapy treatment for sickle cell?
What are the problems with viral vectors? What's been done to address these issues?
What are the proposed ways for obtaining sufficient quantities of gene therapy-modified cells for transplantation?
What determines whether an inserted gene is expressed?

Imren et al., 2004:
This is a research paper that reports original data. To start, read the abstract and introduction to get the context and the general questions to be addressed. Then read the discussion to understand how the findings fit within the context. You now know the start and end point of the paper, so delve into the results section to see what experiments led from one point to the other. Each figure will depict a major finding/set of findings. For each figure, read the associated portion of the text AND the figure legend to determine the specific question addressed, the technique used, and the data generated. Don't bother with the materials and methods section unless you need clarification on particular reagents used. Look up unfamiliar terminology in the textbook and online, but don't get bogged down-- I don't expect you to get every detail as you are initially getting used to reading research papers.
What is the overall goal of this study? How is the approach different from previous studies?
What are lentiviruses? What are their advantages?
What is a self-inactivating lentivirus? Do some detective work online.
How has the beta globin gene in this study been modified to be therapeutic?
What cell types were treated? Into what organisms were the cells transplanted?
Fig 1: what are the components of the lentiviral vector?
The first part of the results section describes a preliminary experiment that is NOT depicted in a figure. What was the purpose?
Fig 2: The blood of treated mice was analyzed. What characteristics were measured? What do all the different lines and symbols depict?
Fig 3: A. How were these cells grown? Were live mice involved?
B. What is FACS analysis? See an explanation from Dr. Campbell's genomics web site. Why was it important that 85% of the treated and cultured cells had glycophorin-A?
C. What do the peaks represent?
D. How were the cells in this FACS analysis generated? Were live mice involved? What do the cell surface markers CD34 and CD45 indicate?
E. What's the difference between this and panel C? (Hint: cell maturity) How does Table 1 related to Fig 3D-E? (Hint: erythroid colonies)
G. What is RT-PCR? What question does it address? What two cell types were compared for expression of the transgene? How are the bottom two panels a control experiment?
Fig 4: See http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=hmg.figgrp.1282 for info on bubble linker PCR. What is the general question addressed in figures 4 and 5? What does the number of bands in each lane of Figure 4 indicate?
Fig 5 and Table 2: How are these data related to Fig 4? What danger of sickle cell gene therapy do these data indicate? Can the researchers move directly into clinical studies in humans?