PCRmax Listeria monocytogenes Instruction Manual

Listeriolysin O (hlyA) gene

Listeria
monocytogenes

PCRmax Ltd qPCR test

TM

150 tests

For general laboratory and research use only

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Listeria monocytogenes is a Gram-positive, rod-shaped bacterium of the Listeriaceae
family. This bacterium is a foodborne pathogen that causes listeriosis. L. moncytogenes
has a circular genome of around 3M nucleotides in length and is motile at room
temperature but not at body temperature.

L. monocytogenes infects humans via the epithelial cells of the intestinal tract after
ingestion of contaminated food. Proteins expressed by the bacteria, including invasins and
internalins, interact with receptors on the host cell mediating direct access to the gut
epithelium. Alternatively, the bacterium may enter the host cell by phagocytosis. After cell
entry, bacterial replication occurs within the mucous membranes of the intestinal lining.
These bacteria specifically target the liver and spleen and translocate to these organs
within around 24 hours of infection. When within the hepatocytes, bacterial replication
increases. In immune-competent people, T-lymphocytes contain the infection within 7 days
and in the majority of cases there is no clinical presentation of the infection. However, in
immuno-compromised people, the infection is not contained by the T-cells and the bacteria
spread to the blood stream causing bacteremia that can lead to listeriosis. Infection of the
blood allows access to the brain and the fetus in pregnant women which can cause
meningoencephalitits and placentitis respectively.
Severe L. monocytogenes infections have high mortality rates between 50% for septicemia
and 80% for neo-natal infections. Antibiotics such as penicillin have been used to
successfully treat some infections.

Introduction to Listeria monocytogenes

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MIN

MAX

The L.monocytogenes primers have been designed for the specific and exclusive in vitro
quantification of all L. monocytogenes and do not detect other Listeria species. The
target sequence (hlyA) has previously been shown to be a good genetic marker for L.
monocytogenes in other real time PCR based studies (Long F, et.al 2008). The primers
and probe sequences in this kit have 100% homology with a broad range of clinically
relevant reference sequences based on a comprehensive bioinformatics analysis.

The PCR Max qPCR Kit for Listeria monocytogenes (L.monocytogenes) genomes is
designed for the in vitro quantification of L.monocytogenes genomes. The kit is designed
to have the broadest detection profile possible whilst remaining specific to the L.
monocytogenes genome.

The primers and probe sequences in this kit have 100% homology with a broad range of L.
monocytogenes sequences based on a comprehensive bioinformatics analysis.

If you require further information, or have a specific question about the detection profile of
this kit then please send an e.mail to [e-mail] and our bioinformatics team will
answer your question.

Specificity

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Kit Contents

• L.monocytogenes specific primer/probe mix (150 reactions BROWN)

FAM labelled

• L.monocytogenes positive control template (for Standard curve RED)

• Internal extraction control primer/probe mix (150 reactions BROWN)

VIC labelled as standard

• Internal extraction control DNA (150 reactions BLUE)

• Endogenous control primer/probe mix (150 reactions BROWN)

FAM labelled

• RNAse/DNAse free water (WHITE)

for resuspension of primer/probe mixes and internal extraction control DNA

• Template preparation buffer (YELLOW)

for resuspension of positive control template and standard curve preparation

Reagents and equipment to be supplied by the user

Real-Time PCR Instrument

DNA extraction kit

This kit designed to work well with all processes that yield high quality DNA with minimal
PCR inhibitors.

Lyophilised 2x qPCR Mastermix

This kit is designed to be compatible with all commercially available Mastermixes that run
with standard cycling conditions.

Pipettors and Tips

Vortex and centrifuge

Thin walled 1.5 ml PCR reaction tubes

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