Cica Geneus Pathogenesis GeneDetection PCR Kit （for Diarrheagenic ）TMDiarrheagenic（DEC） causes abdominal pain, diarrheal disease, andDEC is classiﬁed in seven kinds by the diﬀerence in property and symptom.
This method of this kit is to amplify plural speciﬁc genes by Multiplex PCR at thesame time, and then to analyze agarose gel electrophoresis banding patterns.
This kit can detect ﬁve kinds of representative pathogenesis genes.

Characteristics●Using three kinds of primer mixes, this kit can detect pathogenesis genes
about EHEC, EPEC, ETEC, EIEC, EAEC．
●This kit can clarify a judgement result by digitizing a detection result.
■ Composition（100 rxns）
Reagent■ Work FlowVolumeA

AptaTaq DNA Master（5×Conc.）*１

1.2 mL × 1

B

PCR supplement

1.1 mL × 2

CPrimer mix 1

0.4 mL × 1

D

Primer mix 2

0.4 mL × 1

E

Primer mix 3

0.4 mL × 1

F

Positive control

0.4 mL × 1

GLoading buﬀer（6×Conc.）

1.2 mL × 1

Isolate*1 AptaTaq DNA Master（5×Conc.）is the product of Roche Diagnostics K.K.
*2 CicaGeneus® DNA extraction reagent is sold separately.

Kanto Chemical

DNA Extraction*2

Multiplex PCR
Agarose gelelectrophoresis

Decision

Analyzebanding patterns

Table.1 Kind of detection genes and Amplicon size（bp） Target
Coeﬃcient Amplicon size
Reaction
mixture1
Reaction
mixture2
Reaction
mixture3

1

384

32

132

64

93

16

130

862

2575

4382

Internal control

120

16S rDNA① Read banding patterns（Table.1）from an electrophoreticresult（Fig.1）.

②Using having banding patterns or not, you input in an
exclusive calculating formula and calculate a total levelof the judgment and judge the pathotype. (Table.2)●When 16S rDNA is positive, you can estimate that it is.
●Themay fall oﬀ for a certain invasiveness geneon plasmid.
●This product cannot distinguishfrom EIEC.

Fig.1 Example of the electrophoretic patterns
3：ATCCTM 35401（ETEC）

100：100 bp DNA Ladder
P：Positive control
1：ATCCTM 35150（EHEC）
2：ATCCTM 43887（EPEC）

4：ATCCTM 43893（EIEC）
5：ATCCTM 25922（not DEC）
6〜10：Clinical isolate

Table.2 Total point and Kind of pathonenesis
Ｔｏｔａｌｐｏｉｎｔ Kind of pathogenesis
except EHEC/STEC、EPEC、ETEC、EIEC、EAEC or closely related microbe0■ Result
Sample Number in Fig.1

Category

1EPEC

2EAEC

4〜7EIEC

123456789108〜11ETEC（＋）

Total point97

124409712416

216〜19ETEC（＋）

Decision

EHEC EPEC ETEC EIEC Other EHEC EPEC ETEC ETEC EAEC

24〜27

ETEC
（＋、 ＋）

32〜35

EHEC/STEC
（＋）

64〜67

EHEC/STEC
（＋）

96〜99

EHEC/STEC（＋、Other No.

Mixed infection is doubted＋）

Product Information
Product No.

Product Name08187-96

CicaGeneus Pathogenesis Gene Detection PCR Kit（for Diarreagenic

08178-96

CicaGeneus

TMTM

DNA Extraction Reagent

Multiplex PCR kit series（stored at -20 ℃ 〜 -25 ℃）
Product No.

）

Product Name

Package size

Package size

Sｔｏｒｅd at

100 rxns-20 ℃〜-25 ℃

120 rxns

2 ℃〜8 ℃

Related reagents
Product Name

Product No.

Package size

08180-96 CicaGeneusTM Staph POT KIT

120 rxns

46510-79 10×TBE buﬀer

08187-96 CicaGeneusTM Pseudo POT KIT

50 rxns

46509-79 10×TAE buﬀer

08062-96 CicaGeneusTM Acineto POT KIT

30 rxns

14575-43 Ethidium bromide solution

08179-96 CicaGeneusTM Staph coaglase detection set

50 rxns

01089-23 Agarose KANTO HC

100 g

08081-96 CicaGeneusTM Bovine Mycoplasma HI screening kit

100 rxns

01016-96 AptaTaq DNA Master（5×Conc.）

500 μL

1L
1L
10 mL

KANTO CHEMICAL CO., INC.
REAGENT DIVISION

East Muromachi Mitsui BLDG, 2-1, Nihonbashi Muromachi 2-chome,
Chuo-ku, Tokyo, 103-0022, JAPAN
Telephone+81-3-6214-1092
Telefax+81-3-3241-1053
http://www.kanto.co.jp
E-mail; kanto-61@gms.kanto.co.jp2015. August.