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Unambiguous molecular detections with multiple genetic approach for the complicated chromosome 22q11 deletion syndrome.

Yang C, Huang CH, Cheong ML, Hung KL, Lin LH, Yu YS, Chien CC, Huang HC, Chen CW, Huang CJ - BMC Med. Genet. (2009)

Bottom Line: Changes in genomic copy number were significantly associated with clinical manifestations for the classical criteria of 22q11DS using MPLA and qPCR (p < 0.01).These reduced DNA dosages were also obtained partially using array-CGH and confirmed by qPCR but with some differences in deletion size.These data demonstrate that such multiple genetic approaches are necessary for the unambiguous molecular detection of these types of complicated genomic syndromes.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Genetics, Department of Pediatrics, Taipei Medical University Hospital, Taipei 11031, Taiwan. yeungsann@yahoo.com.tw

ABSTRACT

Background: Chromosome 22q11 deletion syndrome (22q11DS) causes a developmental disorder during the embryonic stage, usually because of hemizygous deletions. The clinical pictures of patients with 22q11DS vary because of polymorphisms: on average, approximately 93% of affected individuals have a de novo deletion of 22q11, and the rest have inherited the same deletion from a parent. Methods using multiple genetic markers are thus important for the accurate detection of these microdeletions.

Methods: We studied 12 babies suspected to carry 22q11DS and 18 age-matched healthy controls from unrelated Taiwanese families. We determined genomic variance using microarray-based comparative genomic hybridization (array-CGH), quantitative real-time polymerase chain reaction (qPCR) and multiplex ligation-dependent probe amplification (MLPA).

Results: Changes in genomic copy number were significantly associated with clinical manifestations for the classical criteria of 22q11DS using MPLA and qPCR (p < 0.01). An identical deletion was shown in three affected infants by MLPA. These reduced DNA dosages were also obtained partially using array-CGH and confirmed by qPCR but with some differences in deletion size.

Conclusion: Both MLPA and qPCR could produce a clearly defined range of deleted genomic DNA, whereas there must be a deleted genome that is not distinguishable using MLPA. These data demonstrate that such multiple genetic approaches are necessary for the unambiguous molecular detection of these types of complicated genomic syndromes.

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Related in: MedlinePlus

DNA dosage profiles determined by quantitative real-time PCR. Histograms of relative DNA dosage for (A) 18 healthy controls and (B) 12 patients suspected to carry 22q11DS. Ratios on each DNA dosage are plotted and normalized with a reference endogenous gene, GAPDH (AY340484). Eight probes are ordered by position on chromosome 22q. Samples retaining both alleles, log2 ratio close to 0; samples with deleted one allele, log2 ratio close to -1.
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Figure 4: DNA dosage profiles determined by quantitative real-time PCR. Histograms of relative DNA dosage for (A) 18 healthy controls and (B) 12 patients suspected to carry 22q11DS. Ratios on each DNA dosage are plotted and normalized with a reference endogenous gene, GAPDH (AY340484). Eight probes are ordered by position on chromosome 22q. Samples retaining both alleles, log2 ratio close to 0; samples with deleted one allele, log2 ratio close to -1.

Mentions: To confirm and clearly define genomic deletions in the proband infants, we applied qPCR using TaqMan probes. Compared with the 18 healthy controls, DNA dosage levels from the 12 patients varied independently (Figure 4). Genomic imbalance was detected in four of these babies (B01, B02, B03 and B12), but no detectable changes were found in the other eight babies or in the healthy controls. Changes in genomic copy numbers were also significantly associated with clinical manifestations (P < 0.01 by Fisher's exact test). As with the MLPA technique, subjects B01, B02 and B12 showed large genomic deletions in 22q11 by qPCR (range of log2 ratios -0.79 to -1.32). However, the size of the deletion in proband B01 was smaller than in B02 and B12. As shown in Figure 4b, an additional haploinsufficiency for probe T3M was detected from probands B02 and B12 (log2 ratios -1.15 and -0.95, respectively) but not from subject B01 (log2 ratio +0.22). This proband carried a 2.38 Mb deletion (from probes A3M to T3M) whereas subjects B02 and B12 each had a 2.27 Mb deletion (from probes A3M to LZTR1-2). Moreover, subject B03 showed very low DNA dosage (log2 ratio -3.06) by qPCR, using probe CO3M.


Unambiguous molecular detections with multiple genetic approach for the complicated chromosome 22q11 deletion syndrome.

Yang C, Huang CH, Cheong ML, Hung KL, Lin LH, Yu YS, Chien CC, Huang HC, Chen CW, Huang CJ - BMC Med. Genet. (2009)

DNA dosage profiles determined by quantitative real-time PCR. Histograms of relative DNA dosage for (A) 18 healthy controls and (B) 12 patients suspected to carry 22q11DS. Ratios on each DNA dosage are plotted and normalized with a reference endogenous gene, GAPDH (AY340484). Eight probes are ordered by position on chromosome 22q. Samples retaining both alleles, log2 ratio close to 0; samples with deleted one allele, log2 ratio close to -1.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2656481&req=5

Figure 4: DNA dosage profiles determined by quantitative real-time PCR. Histograms of relative DNA dosage for (A) 18 healthy controls and (B) 12 patients suspected to carry 22q11DS. Ratios on each DNA dosage are plotted and normalized with a reference endogenous gene, GAPDH (AY340484). Eight probes are ordered by position on chromosome 22q. Samples retaining both alleles, log2 ratio close to 0; samples with deleted one allele, log2 ratio close to -1.
Mentions: To confirm and clearly define genomic deletions in the proband infants, we applied qPCR using TaqMan probes. Compared with the 18 healthy controls, DNA dosage levels from the 12 patients varied independently (Figure 4). Genomic imbalance was detected in four of these babies (B01, B02, B03 and B12), but no detectable changes were found in the other eight babies or in the healthy controls. Changes in genomic copy numbers were also significantly associated with clinical manifestations (P < 0.01 by Fisher's exact test). As with the MLPA technique, subjects B01, B02 and B12 showed large genomic deletions in 22q11 by qPCR (range of log2 ratios -0.79 to -1.32). However, the size of the deletion in proband B01 was smaller than in B02 and B12. As shown in Figure 4b, an additional haploinsufficiency for probe T3M was detected from probands B02 and B12 (log2 ratios -1.15 and -0.95, respectively) but not from subject B01 (log2 ratio +0.22). This proband carried a 2.38 Mb deletion (from probes A3M to T3M) whereas subjects B02 and B12 each had a 2.27 Mb deletion (from probes A3M to LZTR1-2). Moreover, subject B03 showed very low DNA dosage (log2 ratio -3.06) by qPCR, using probe CO3M.

Bottom Line: Changes in genomic copy number were significantly associated with clinical manifestations for the classical criteria of 22q11DS using MPLA and qPCR (p < 0.01).These reduced DNA dosages were also obtained partially using array-CGH and confirmed by qPCR but with some differences in deletion size.These data demonstrate that such multiple genetic approaches are necessary for the unambiguous molecular detection of these types of complicated genomic syndromes.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Genetics, Department of Pediatrics, Taipei Medical University Hospital, Taipei 11031, Taiwan. yeungsann@yahoo.com.tw

ABSTRACT

Background: Chromosome 22q11 deletion syndrome (22q11DS) causes a developmental disorder during the embryonic stage, usually because of hemizygous deletions. The clinical pictures of patients with 22q11DS vary because of polymorphisms: on average, approximately 93% of affected individuals have a de novo deletion of 22q11, and the rest have inherited the same deletion from a parent. Methods using multiple genetic markers are thus important for the accurate detection of these microdeletions.

Methods: We studied 12 babies suspected to carry 22q11DS and 18 age-matched healthy controls from unrelated Taiwanese families. We determined genomic variance using microarray-based comparative genomic hybridization (array-CGH), quantitative real-time polymerase chain reaction (qPCR) and multiplex ligation-dependent probe amplification (MLPA).

Results: Changes in genomic copy number were significantly associated with clinical manifestations for the classical criteria of 22q11DS using MPLA and qPCR (p < 0.01). An identical deletion was shown in three affected infants by MLPA. These reduced DNA dosages were also obtained partially using array-CGH and confirmed by qPCR but with some differences in deletion size.

Conclusion: Both MLPA and qPCR could produce a clearly defined range of deleted genomic DNA, whereas there must be a deleted genome that is not distinguishable using MLPA. These data demonstrate that such multiple genetic approaches are necessary for the unambiguous molecular detection of these types of complicated genomic syndromes.

Show MeSH
Related in: MedlinePlus