Abstract
Several species of helicobacter have been isolated from laboratory mice, including H. bilis, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius, which appear to be the most common. The most widely used published method for molecular detection of these agents is PCR amplification of a conserved region of 16S rRNA, but differential speciation requires restriction enzyme digestion of the amplicons. This study was undertaken to determine PCR conditions that would simultaneously and specifically identify each of the five common species without restriction enzyme analyses. First, we designed novel and specific PCR primers for H. bills, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius, using sequences from the heterologous regions of 16S rRNA. Because of comigration of amplified products, we next identified P17, an H. bilis-specific protein; P25, an H. hepaticus-specific protein; and P30, an H. muridarum-specific protein by screening genomic DNA expression libraries of each species. Primers were designed from these three genes, plus newly designed, species-specific 16S rRNA primers for H. rodentium and H. typhlonius that could be utilized for a five-plex PCR. The sizes of the amplicons from H. bilis, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius were 435, 705, 807, 191, and 122 bp, respectively, allowing simultaneous detection and effective discrimination among species.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 531-536 |
| Number of pages | 6 |
| Journal | Clinical and Diagnostic Laboratory Immunology |
| Volume | 12 |
| Issue number | 4 |
| DOIs | |
| State | Published - Apr 2005 |
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ASJC Scopus subject areas
- Microbiology (medical)
- Immunology and Allergy
- Clinical Biochemistry
- Immunology
Cite this
Differential detection of five mouse-infecting helicobacter species by multiplex PCR. / Feng, Sunlian; Ku, Karin; Hodzic, Emir; Lorenzana, Edward; Freet, Kim; Barthold, Stephen W.
In: Clinical and Diagnostic Laboratory Immunology, Vol. 12, No. 4, 04.2005, p. 531-536.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Differential detection of five mouse-infecting helicobacter species by multiplex PCR
AU - Feng, Sunlian
AU - Ku, Karin
AU - Hodzic, Emir
AU - Lorenzana, Edward
AU - Freet, Kim
AU - Barthold, Stephen W
PY - 2005/4
Y1 - 2005/4
N2 - Several species of helicobacter have been isolated from laboratory mice, including H. bilis, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius, which appear to be the most common. The most widely used published method for molecular detection of these agents is PCR amplification of a conserved region of 16S rRNA, but differential speciation requires restriction enzyme digestion of the amplicons. This study was undertaken to determine PCR conditions that would simultaneously and specifically identify each of the five common species without restriction enzyme analyses. First, we designed novel and specific PCR primers for H. bills, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius, using sequences from the heterologous regions of 16S rRNA. Because of comigration of amplified products, we next identified P17, an H. bilis-specific protein; P25, an H. hepaticus-specific protein; and P30, an H. muridarum-specific protein by screening genomic DNA expression libraries of each species. Primers were designed from these three genes, plus newly designed, species-specific 16S rRNA primers for H. rodentium and H. typhlonius that could be utilized for a five-plex PCR. The sizes of the amplicons from H. bilis, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius were 435, 705, 807, 191, and 122 bp, respectively, allowing simultaneous detection and effective discrimination among species.
AB - Several species of helicobacter have been isolated from laboratory mice, including H. bilis, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius, which appear to be the most common. The most widely used published method for molecular detection of these agents is PCR amplification of a conserved region of 16S rRNA, but differential speciation requires restriction enzyme digestion of the amplicons. This study was undertaken to determine PCR conditions that would simultaneously and specifically identify each of the five common species without restriction enzyme analyses. First, we designed novel and specific PCR primers for H. bills, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius, using sequences from the heterologous regions of 16S rRNA. Because of comigration of amplified products, we next identified P17, an H. bilis-specific protein; P25, an H. hepaticus-specific protein; and P30, an H. muridarum-specific protein by screening genomic DNA expression libraries of each species. Primers were designed from these three genes, plus newly designed, species-specific 16S rRNA primers for H. rodentium and H. typhlonius that could be utilized for a five-plex PCR. The sizes of the amplicons from H. bilis, H. hepaticus, H. muridarum, H. rodentium, and H. typhlonius were 435, 705, 807, 191, and 122 bp, respectively, allowing simultaneous detection and effective discrimination among species.
UR - http://www.scopus.com/inward/record.url?scp=22844444194&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=22844444194&partnerID=8YFLogxK
U2 - 10.1128/CDLI.12.4.531-536.2005
DO - 10.1128/CDLI.12.4.531-536.2005
M3 - Article
C2 - 15817762
AN - SCOPUS:22844444194
VL - 12
SP - 531
EP - 536
JO - Clinical and Vaccine Immunology
JF - Clinical and Vaccine Immunology
SN - 1556-6811
IS - 4
ER -