PCRmax Clostridium difficile (toxin B) Instruction Manual

Toxin B (toxB) gene

Clostridium difficile
(toxin B)

PCRmax Ltd qPCR test

TM

150 tests

For general laboratory and research use only

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Clostridium difficile is a motile, rod-shaped, Gram-positive bacterium of the Clostridiaceae
family. The genome of this bacterium is around 4.3 Mb arranged as a single, circular
chromosome with a plasmid. A number of genes are responsible for virulence, host
interaction and resistance. Infection with this bacterium is known to be the leading cause of
antibiotic-associated diarrhoea, especially in relation to nosocomial infections.

C. difficile is not a major component of the natural flora found in the human gut but
treatment with broad-spectrum antibiotics allows the bacterium to colonise where other
bacterial species have been removed. Transmission of C. difficile is by the faecal-oral
route. Under stress, such as exposure to disinfectants, the bacterium forms spores which
are capable of survival under extreme conditions. When conditions become more
favourable, such as when the bacterium reaches the intestine after ingestion, the spore
can convert back to the active bacterial form. Once the bacterium has colonized the
intestine, the production of toxins is upregulated. The bacterium produces two toxins; Toxin
A, an enterotoxin and toxin B, a cytotoxin. Toxin B is found in all C. difficile strains whereas
toxin A is found in only 70% of strains. The toxins work to disrupt tight junctions of the
intestinal epithelial cells resulting in inflammation and increased permeability of the
intestine.

Infection with C. difficile results in symptoms such as fever, abdominal pains and diarrhoea
which can be due to pseudomembranous colitis (PMC), an inflammation of the colon. The
infection can be treated with antibiotics although resistance to some antibiotics has been
shown.

Introduction to Clostridium difficile (toxin B)

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MIN

MAX

The PCR Max qPCR Kit for Clostridium difficile (toxin B) (C.difficile_toxB) genomes is
designed for the in vitro quantification of C.difficile_toxB genomes. The kit is designed to
have the broadest detection profile possible whilst remaining specific to the C.difficile_toxB
genome.

The primers and probe sequences in this kit have 100% homology with a broad range of
C.difficile_toxB 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

• C.difficile_toxB specific primer/probe mix (150 reactions BROWN)

FAM labelled

• C.difficile_toxB 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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