Term
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Definition
| Multiplex Ligation Probe Dependent Amplification |
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Term
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Definition
| Each probe consists of 2 pieces: 1) synthetic oligo (50-60bp) consisting of PCR primer sequence and hybridization sequence. 2) M13-derived oligonucleotide (60-450bp) consisting of PCR primer sequence, stuffer sequence and hybridization sequence. |
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Term
| Denaturation and Hybridization |
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Definition
MLPA probemix added to denatured genomic DNA.
Two parts of each probe hybridize to adjacent target sequences.
Hybridization will only occur if probe sequence is complementary to the template. |
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Term
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Definition
| If both adjacent probe halves are complementary to template, probes are ligated by a thermostable ligase. |
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Term
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Definition
| When next denaturation step occurs, ligated probe remains together, and is a template for PCR amplification using the universal primer set. |
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Term
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Definition
By comparison to control sample, it is possible to detect deletions/duplications based on differences in relative peak heights.
Alternative: use specialty programs to normalize data: adjusts the peak intensities within individual samples to the same scale for comparative purposes. |
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Term
| Methylation Sensitive MLPA |
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Definition
methylation present = sequence not digested = MLPA amplification can occur
no methylation present = sequence digest = MLPA amplification not possible. |
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Term
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Definition
Normal individual: 2 copies of Hha1 site: undigested (MLPA): normal peak height. Digested (MS-MLPA): 1/2 normal peak height.
PWS (paternal Ch15 del): only maternal copy of SNRPN=methylated. Undigested (MLPA): 1/2 normal peak height. Digested (MS-MLPA): 1/2 normal peak height.
PWS (maternal UPD). Undigested: normal peak height. Digested (MS-MLPA): normal peak height.
AS (maternal Ch15 del): Undigested: 1/2 normal peak height. Digested (MS-MLPA): no peak.
AS (paternal UPD): Undigested: normal peak height. Digested (MS-MLPA): not methylated therefore digested no peak |
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