PCRmax H10N8 Instruction Manual

Hemagglutinin (HA) gene &
Neuraminidase (NA) gene

H10N8

PCRmax Ltd qPCR test

TM

150 tests

For general laboratory and research use only

1

Quantification of H10N8 genomes

Advanced kit handbook HB10.01.09

Published Date: 13/10/2016

Avian Influenza (‘avian flu’ or ‘bird flu’) strains all belong to the Influenza A virus which are
generally adapted to birds but not exclusive to them.

Of particular concern to humans are ones which are infectious to both humans and birds.
Specific strains such as H1N1 have been the subject of much media concern and
speculation over pandemics and its widespread transmission globally.

Only some strains of avian influenza are pathogenic in humans typically H5N1, H7N3,
H7N7 and H7N9 but now H10N8 has proven fatal in China as of Dec 2013.

Influenza type A viruses are 80–120 nanometers in diameter and usually roughly spherical,
made up of a viral envelope containing two main types of proteins, wrapped around a
central core.

The two large proteins found on the outside of viral particles arehemagglutinin
(HA)andneuraminidase (NA). HA is a protein that mediates binding of the virion to target
cells and entry of the viral genome into the target cell, while NA is involved in the release of
progeny virions from infected cells

Influenza type A viruses are categorized into subtypes based on the type of these two
proteins on the surface of the viral envelope.

The central core of a virion contains the viral genome and other viral proteins that package
and protect the genetic material
The entire Influenza A virus genome is ~13,588 bases long and is contained on8RNA
segments that code for11proteins.

Introduction to H10N8

2

Quantification of H10N8 genomes

Advanced kit handbook HB10.01.09

Published Date: 13/10/2016

Specificity

The PCRmax™ qPCR Kit for H10N8 (H10N8) genomes is designed for the in vitro
quantification of H10N8 genomes. The kit is designed to have the broadest detection
profile possible whilst remaining specific to the H10N8 genome.
The primers and probe sequences in this kit have 100% homology with a broad range of
H10N8 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.

3

Quantification of H10N8 genomes

Advanced kit handbook HB10.01.09

Published Date: 13/10/2016

Kit Contents

• H10N8 specific primer/probe mix (150 reactions BROWN)

FAM labelled

• H10N8 positive control template (for Standard curve RED)

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

VIC labelled as standard

• Internal extraction control RNA (150 reactions BLUE)

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

FAM labelled

• H10N8/Internal extraction control/endogenous control RT primer mix (150
reactions GREEN)

Required for two step protocol only

• RNAse/DNAse free water (WHITE)

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

• 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

RNA extraction kit

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

Lyophilised OneStep 2x RT-qPCR Mastermix

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

Pipettors and Tips

Vortex and centrifuge

Thin walled 1.5 ml PCR reaction tubes

4

Quantification of H10N8 genomes

Advanced kit handbook HB10.01.09

Published Date: 13/10/2016