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Chapter 30 : Molecular Diagnosis of Chronic Granulomatous Disease

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Abstract:

This chapter focuses on defining the chronic granulomatous disease (CGD) subtype and identifying the specific genetic defect. Before genetic analysis of CGD patients is performed, it is advisable to identify the affected protein component to pinpoint the defective gene. The molecular defects that cause the X91, A22, and A67 forms of CGD are highly heterogeneous in nature, with many of them being family specific. In addition to confirming the CGD subtype, the identification of specific mutations can provide important information for detecting or confirming carriers among patients’ family members and for prenatal diagnosis in affected families. Once genomic DNA has been isolated, single-strand conformational polymorphism (SSCP) analysis can be used initially to examine the 13 exons of the gene, in order to identify the region likely to contain a mutation. More than 90% of all A47 CGD patients analyzed to date are homozygous for the deletion of GT from the normal GTGT at the beginning of exon 2 (ΔGT/ΔGT genotype).

Citation: Heyworth P, Curnutte J. 2006. Molecular Diagnosis of Chronic Granulomatous Disease, p 262-271. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch30

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Integral Membrane Proteins
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Chronic Granulomatous Disease
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Plasma Membrane
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Figures

Image of FIGURE 1
FIGURE 1

Reduced-minus-oxidized difference spectra of Triton X-100 extracts of neutrophils from a patient with X-linked CGD, his mother (carrier), and a normal control. The absorbance bands at 427, 530 and 559 nm, characteristic of flav-ocytochrome , are indicated (arrows). The broad absorbance band around 470 nm is due to myeloperoxidase. The spectra have been manually offset for clarity.

Citation: Heyworth P, Curnutte J. 2006. Molecular Diagnosis of Chronic Granulomatous Disease, p 262-271. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch30
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Image of FIGURE 2
FIGURE 2

SSCP analysis of exons 1 through 7 of the gene. After electrophoresis of amplified and denatured DNA, the gel was stained with SYBR Green II. In each bracketed pair of lanes, the patient sample is on the left and the normal control is on the right. In the analysis shown, exon 1 of the patient contained an aberrant band (arrow); for all the other exons, the control and patient single-strand bands were the same. A four-nucleotide insertion was subsequently identified in exon 1 of this patient’s gene. Nondenatured, double-stranded DNA appears at the bottom of the gel.

Citation: Heyworth P, Curnutte J. 2006. Molecular Diagnosis of Chronic Granulomatous Disease, p 262-271. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch30
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References

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Tables

Generic image for table
TABLE 1

Oligonucleotide primers for PCR amplification of

Citation: Heyworth P, Curnutte J. 2006. Molecular Diagnosis of Chronic Granulomatous Disease, p 262-271. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch30
Generic image for table
TABLE 2

Oligonucleotide primers for PCR amplification and sequencing of

Citation: Heyworth P, Curnutte J. 2006. Molecular Diagnosis of Chronic Granulomatous Disease, p 262-271. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch30
Generic image for table
TABLE 3

Oligonucleotide primers for PCR amplification and sequencing of exon 2 of 1

Citation: Heyworth P, Curnutte J. 2006. Molecular Diagnosis of Chronic Granulomatous Disease, p 262-271. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch30
Generic image for table
TABLE 4

Oligonucleotide primers for PCR amplification and sequencing of

Citation: Heyworth P, Curnutte J. 2006. Molecular Diagnosis of Chronic Granulomatous Disease, p 262-271. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch30

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