MutaPLATE® Lactase PCR Assay Kit

$840.00

The Eagle Biosciences MutaPLATE® Lactase real-time PCR Assay kit is a molecular biological test for analysis of -13910 T/C – polymorphism in the regulatory region of the lactase-phlorizin-hydrolase gene (LPH) in open real-time PCR systems e. g. RotorGene (Corbett Research), MX-3000P (Stratagene), ECO (Amplifa) or SmartCycler (Cepheid). Only the clinical relevant C/C – genotype causes a primary lactase deficiency often leading to (genetic) lactose intolerance. The Eagle Biosciences MutaPLATE® Lactase is for research use only and is not intended for diagnostic or therapeutic procedures.

SKU: KF1907132 Categories: ,

MutaPLATE® Lactase PCR Assay Kit

For Research Use Only

Size: 32 tests
Method: PCR
Sample Type: DNA (e.g.whole blood, cheek swab)
Sample Size: 200 µL

Product manufactured by Immundiagnostik

Additional Information

Assay Background

Patients with lactose intolerance can not digest milk sugar and suffer after ingestion of milk-products from dyspepsia, nausea, and bellyache. Further symptoms like vertigo, sleep disorders, acne or depressions can also be triggered by lactose intolerance. A Therapy for affected persons is very simple and can be done by lactose-free diet. In Germany, about 15 million people are affected by primary lactase deficiency [1].

The main reason for lactose intolerance is a genetically based deficiency of the enzyme lactase-phlorizin hydrolase (LPH), which is responsible for the disassembly of milk sugar. This widely distributed genetic disorder is a T/C polymorphism located at position –13910 in the regulatory region of this gene [2]. Person homozygous for C/C-genotype are consequently deficient for enzyme lactase and posses higher risk for lactose intolerance. These results are in excellent accordance with results obtained by the lactose hydrogen breath test for the diagnosis of lactase non-persistence [3]. Nevertheless, not all C/C-carriers must show typical symptoms because a fall short of the individual level is necessary.

Furthermore, in some cases, lactose intolerance can be due to secondary causes like mal-resorption problems (e. g. Morbus Crohn patients), infections or chemotherapy [4].

In babyhood and infancy, the lactase production is very high but it decreases with higher age resulting in the manifestation of primary lactase deficiency. Also, a North-/ South gradient is visible: In Scandinavia, the homozygous C/C-constellation is very rare whereas in Germany prevalence is about 15-20%. In Southern European countries up to 30% of all adults carry the C-alleles homozygous.

Patients suffering from lactose intolerance have also a higher risk of osteoporosis due to the reduced calcium-intake via milk products [5]. In consequence, the C/C-genotype associated with primary lactose intolerance is a genetic risk factor for bone fractures for elderly people [6].

Assay Principle

The Eagle Biosciences MutaPLATE® Lactase real-time PCR Kit contains specific primers and additional material for the detection of the T/C (-13910) polymorphism in the regulatory region of the lactase-phlorizin-hydrolase gene. The variable area of the regulatory region from lactase gene is amplified by PCR using genomic DNA-template. The specific primers used in the kit flank the variable area of the lactase gene (LCT) and generate an amplification of 222 bp.

The standard PCR contains also sequence-specific oligonucleotides marked with fluorescence dye. This specific hybridization probe binds within the amplification product including the single nucleotide polymorphism (SNP) of target-DNA. Due to this, a fluorescence signal is generated (after excitation with 530 nm or if limited by an instrument with 470 nm) and detected at 610 nm through the optical unit of the real-time PCR instrument.

Genotyping is performed by subsequent melting curve analysis of arisen amplificates leading to unequivocal identification of C/C-genotype associated with lactose intolerance and respectively the clinical unobtrusive CT- and T/T-variants. This is due to the different melting points of the complexes formed by DNA template and “SNP-probes”. The included “SNP-probe” is 100% homologous to the C-allele. Therefore the hybridization probe needs a higher temperature for complex-dissociation from C-allele than from the T-allele (containing a mismatch destabilizing the complex). Consequently, samples with heterozygous genotype generate both peaks at different temperatures during the melting curve analysis.

Manual

Product Manual


Publications

Refrences

  1. Buning C, Ockenga J, Kruger S, Jurga J, Baier P, Dignass A, Vogel A, Strassburg C, Weltrich R, Genschel J, Lochs H, Schmitt H. (2003) The C/C(-13910) and G/G (-22018) genotypes for adult-type hypolactasia are not associated with IBD. Scand J Gastroenterol 38: 538-542.
  2. Enattah NS, Sahi T, Savilahti E, Terwilliger JD, Peltonen L, Jarvela I. (2002). Identification of a variant associated with adult-type hypolactasia. Nat Genet 30: 233-237.
  3. Högenauer C, Hammer HF, Mellitzer K, Renner W, Krejs GJ, Toplak H (2005). Evaluation of a new DNA test compared with the lactose hydrogen breath test for the diagnosis fo lactase non-persistence. Eur J Gastroenterol Hepatol 17: 371-376.
  4. Sibley E (2004) Genetic Variation/ Lactose Intolerance. Detection Methods/ Clinical Implications. Am J Pharmacogenomics 4 (4): 239-245.
  5. Obermeyer-Pietsch BM, Bonelli CM, Walter DE, Kuhn RJ, Fahrleitner-PA, Berghold A, Goessler W, Stepan V, Dobnig H, Leb G, Renner W (2004) Genetic predisposition for adult lactose intolerance and relation to diet, bone density, bone fractures. J Bone a. Mineral Research 19 (1): 42-47.
  6. Ennattah NS, Sulkava R, Halonen P, Kontula K, Järvelä I (2005) Genetic variant of lactase-persistent C/T- -13910 is associated with bone fractures in very old age. JAGS 53: 79-82.