Array comparative genomic hybridization
Encyclopedia
Array-comparative genomic hybridization (also CMA, Chromosomal microarray analysis, microarray-based comparative genomic hybridization, array CGH, a-CGH, aCGH, or virtual karyotype
) is a technique to detect genomic copy number variations at a higher resolution level than chromosome-based comparative genomic hybridization
(CGH).
from a test sample and normal reference sample are labelled differentially, using different fluorophores, and hybridized to several thousand probes. The probes are derived from most of the known genes
and non-coding regions of the genome
, printed on a glass slide.
The fluorescence
intensity of the test and of the reference DNA is then measured, to calculate the ratio between them and subsequently the copy number changes for a particular location in the genome.
s (SV) at resolution of 200 bp . This method allows one to identify new recurrent chromosome changes such as
in disease conditions such as cancer
and birth defects due to chromosome aberrations.
Virtual Karyotype
Virtual karyotype detects genomic copy number variations at a higher resolution level than conventional karyotyping or chromosome-based comparative genomic hybridization .-Background:...
) is a technique to detect genomic copy number variations at a higher resolution level than chromosome-based comparative genomic hybridization
Comparative genomic hybridization
Comparative genomic hybridization or Chromosomal Microarray Analysis is a molecular-cytogenetic method for the analysis of copy number changes in the DNA content of a given subject's DNA and often in tumor cells....
(CGH).
Process
DNADNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
from a test sample and normal reference sample are labelled differentially, using different fluorophores, and hybridized to several thousand probes. The probes are derived from most of the known genes
Gênes
Gênes is the name of a département of the First French Empire in present Italy, named after the city of Genoa. It was formed in 1805, when Napoleon Bonaparte occupied the Republic of Genoa. Its capital was Genoa, and it was divided in the arrondissements of Genoa, Bobbio, Novi Ligure, Tortona and...
and non-coding regions of the genome
Genome
In modern molecular biology and genetics, the genome is the entirety of an organism's hereditary information. It is encoded either in DNA or, for many types of virus, in RNA. The genome includes both the genes and the non-coding sequences of the DNA/RNA....
, printed on a glass slide.
The fluorescence
Fluorescence
Fluorescence is the emission of light by a substance that has absorbed light or other electromagnetic radiation of a different wavelength. It is a form of luminescence. In most cases, emitted light has a longer wavelength, and therefore lower energy, than the absorbed radiation...
intensity of the test and of the reference DNA is then measured, to calculate the ratio between them and subsequently the copy number changes for a particular location in the genome.
Efficiency
Using this method, copy number changes at a level of 5–10 kilobases of DNA sequences can be detected. Today even high-resolution CGH (HR-CGH) arrays are accurate to detect structural variationStructural variation
Structural variation is the variation in structure of an organism's chromosome. It consists of many kinds of variation in the genome of one species, and usually includes microscopic and submicroscopic types, such as deletions, duplications, copy-number variants, insertions, inversions and...
s (SV) at resolution of 200 bp . This method allows one to identify new recurrent chromosome changes such as
- microdeletions
- duplications
in disease conditions such as cancer
Cancer
Cancer , known medically as a malignant neoplasm, is a large group of different diseases, all involving unregulated cell growth. In cancer, cells divide and grow uncontrollably, forming malignant tumors, and invade nearby parts of the body. The cancer may also spread to more distant parts of the...
and birth defects due to chromosome aberrations.
Technical considerations
There are several requirements that are dependent on the application of aCGH:- Complexity. Measurement becomes difficult in larger organisms because of decreasing partial concentrations of each portion of the sequence that is involved in the hybridization to the array element as the size of the genomes increase. This issue may be addressed by increasing the threshold in which one detects only larger increases in copy number of DNA extracted from cells, but this comes at the cost of increasing failure to detect low level gains and losses.
- Samples. Tissue specimens may contain heterogeneous cell populations, which may further decrease the ability to detect copy number change in genes in the aberrant tumor cells because the population may contain normal cells. Furthermore, the use of tissue from clinical specimens severely limits the amount of high quality DNA available for analysis. After isolation of this small DNA amounts, the genomic DNA has to be quantified and assessed for purity and integrity prior to labelling. The labelling efficiency (Frequency of Incorporation, FOI) has to be determined prior to hybridization to adjust the amounts of labelled DNA between patient and reference to an equal amount. Therefore, a cuvetteless small volume spectrophotometer (like the NanoPhotometerTMQuantification of nucleic acidsIn molecular biology, quantitation of nucleic acids is commonly performed to determine the average concentrations of DNA or RNA present in a mixture, as well as their purity. Reactions that use nucleic acids often require particular amounts and purity for optimum performance...
) for precise DNA quantification and quality control is absolute mandatory. - Error tolerance. If the investigator is set to obtain a generalized description of aberrations that may occur in a set of samples, then errors in the detection may not be critical. However, the margin for error is drastically narrowed in a clinical setting, where an individual specimen is used to obtain specific information.