CMA was performed using peripheral blood and examined with Cytoscan HD microarray (Affymetrix, Santa Clara, CA). This array consists of 2,696,550 oligonucleotide probes, including 1,953,246 distinctive non-polymorphic oligonucleotide probes, and 743,304 single nucleotide polymorphism probes. Genomic DNA was extracted and purified from whole blood sample using Gentra Puregene Blood Kit (Qiagen Inc., Valencia, CA). Procedures for DNA digestion, adapter ligation, polymerase chain reaction (PCR), amplicon DNA fragmentation, labeling, and hybridization of the arrays were performed according to manufacturer’s instructions (Affymetrix, Santa Clara, CA). Results were investigated using the Chromosome Analysis Suite (ChAS; Affymetrix, Santa Clara, CA). The settings for smallest copy number variation (CNV) regions in ChAS were 25 kb and 25 probes for losses, and 50 kb and 50 probes for gains. Genomic linear positions are given relative to NCBI build 37 (hg19) [12 (link)]. Results were interpreted based on published literature, publicly available databases, and by investigating gene content following practice guidelines [13 (link),14 (link)].
Protocol detail
Chromosomal Microarray Analysis of Blood
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Blood
Chromosome
Copy number variation
Digestion
Dna fragmentation
Gene
Genomic
Hybridization
Ligation
Microarray
Oligonucleotide probes
Polymerase chain reaction
Polymorphic
Single nucleotide polymorphism
Corresponding Organization : The University of Texas Medical Branch at Galveston
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Variable analysis
independent variables
- Extraction and purification of genomic DNA from whole blood sample using Gentra Puregene Blood Kit (Qiagen Inc., Valencia, CA)
- DNA digestion, adapter ligation, polymerase chain reaction (PCR), amplicon DNA fragmentation, labeling, and hybridization of the arrays performed according to manufacturer's instructions (Affymetrix, Santa Clara, CA)
- Copy number variation (CNV) regions detected using the Chromosome Analysis Suite (ChAS; Affymetrix, Santa Clara, CA)
- Settings for smallest copy number variation (CNV) regions in ChAS: 25 kb and 25 probes for losses, and 50 kb and 50 probes for gains
- Genomic linear positions given relative to NCBI build 37 (hg19)
- No positive or negative controls were explicitly mentioned in the provided information.
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