f. nucleatum subsp. vincentii jcm11023 t (BioResource International Inc)

BioResource International Inc f. nucleatum subsp. vincentii jcm11023 t

Schematic gene arrangement and positions of primers designed for the amplification of F. <t>nucleatum</t> CRISPR–Cas system. Cas gene, CRISPR region, conserved genes, and non-conserved genes are in pink, yellow, sky blue, and white, respectively. Small orange arrows indicate tRNA genes, and green arrows indicate rRNA genes. The blue vertical line in conserved genes indicates the primer positions that amplify the peripheral region of the CRISPR–Cas system (blue horizontal line). The red vertical line in Cas gene indicates the forward primer position that amplifies the CRISPR region; the reverse primer for this region is common to the reverse primer for peripheral region of the CRISPR–Cas system. If there is a CRISPR region, the red horizontal line region is amplified. Representative strains are not type strains.

F. Nucleatum Subsp. Vincentii Jcm11023 T, supplied by BioResource International Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Schematic gene arrangement and positions of primers designed for the amplification of F. nucleatum CRISPR–Cas system. Cas gene, CRISPR region, conserved genes, and non-conserved genes are in pink, yellow, sky blue, and white, respectively. Small orange arrows indicate tRNA genes, and green arrows indicate rRNA genes. The blue vertical line in conserved genes indicates the primer positions that amplify the peripheral region of the CRISPR–Cas system (blue horizontal line). The red vertical line in Cas gene indicates the forward primer position that amplifies the CRISPR region; the reverse primer for this region is common to the reverse primer for peripheral region of the CRISPR–Cas system. If there is a CRISPR region, the red horizontal line region is amplified. Representative strains are not type strains.

Journal: Microbiology Spectrum

Article Title: Strain-level detection of Fusobacterium nucleatum in colorectal cancer specimens by targeting the CRISPR–Cas region

doi: 10.1128/spectrum.05123-22

Figure Lengend Snippet: Schematic gene arrangement and positions of primers designed for the amplification of F. nucleatum CRISPR–Cas system. Cas gene, CRISPR region, conserved genes, and non-conserved genes are in pink, yellow, sky blue, and white, respectively. Small orange arrows indicate tRNA genes, and green arrows indicate rRNA genes. The blue vertical line in conserved genes indicates the primer positions that amplify the peripheral region of the CRISPR–Cas system (blue horizontal line). The red vertical line in Cas gene indicates the forward primer position that amplifies the CRISPR region; the reverse primer for this region is common to the reverse primer for peripheral region of the CRISPR–Cas system. If there is a CRISPR region, the red horizontal line region is amplified. Representative strains are not type strains.

Article Snippet: Type strains, F. nucleatum subsp. animalis JCM11025 T , F. nucleatum subsp. nucleatum JCM8532 T , F. nucleatum subsp. polymorphum JCM12990 T , and F. nucleatum subsp. vincentii JCM11023 T , were purchased from the Japan Collection of Microorganisms of RIKEN BioResource Research Center.

Techniques: Amplification, CRISPR

Amplification of the targeted CRISPR region of F. nucleatum -type strains by F. nucleatum -strain genotyping PCR. ( a ) Electrophoresis pattern of PCR products of F. nucleatum -type strains. M, DNA ladder marker; an, F. nucleatum subsp. animalis ; nu, F. nucleatum subsp. nucleatum ; po, F. nucleatum subsp. polymorphum ; vi, F. nucleatum subsp. vincentii ; NC, negative control. The size of the marker fragments, from top to bottom, was 20 , 10, 7.0, 5.0 , 4.0, 3.0, 2.0 , 1.5, 1.3, 1.0 , 0.7, 0.5 , 0.4, 0.3, 0.2, and 0.1 kbp; the intensified fragments are underlined. ( b ) DNA sequences of the 5′- and 3′-ends of PCR amplicon of F. nucleatum subsp. animalis Type I-B1 [yellow square in ( a )] are shown and were used for the analysis. Repeat sequences are detected by CRISPRCasFinder and written in red. DNA sequences of other PCR amplicons are in .

Journal: Microbiology Spectrum

Article Title: Strain-level detection of Fusobacterium nucleatum in colorectal cancer specimens by targeting the CRISPR–Cas region

doi: 10.1128/spectrum.05123-22

Figure Lengend Snippet: Amplification of the targeted CRISPR region of F. nucleatum -type strains by F. nucleatum -strain genotyping PCR. ( a ) Electrophoresis pattern of PCR products of F. nucleatum -type strains. M, DNA ladder marker; an, F. nucleatum subsp. animalis ; nu, F. nucleatum subsp. nucleatum ; po, F. nucleatum subsp. polymorphum ; vi, F. nucleatum subsp. vincentii ; NC, negative control. The size of the marker fragments, from top to bottom, was 20 , 10, 7.0, 5.0 , 4.0, 3.0, 2.0 , 1.5, 1.3, 1.0 , 0.7, 0.5 , 0.4, 0.3, 0.2, and 0.1 kbp; the intensified fragments are underlined. ( b ) DNA sequences of the 5′- and 3′-ends of PCR amplicon of F. nucleatum subsp. animalis Type I-B1 [yellow square in ( a )] are shown and were used for the analysis. Repeat sequences are detected by CRISPRCasFinder and written in red. DNA sequences of other PCR amplicons are in .

Techniques: Amplification, CRISPR, Electrophoresis, Marker, Negative Control

Strain-level discrimination of F. nucleatum isolated from saliva of CRC patients by CRISPR-associated region-targeted PCR. Representative isolates of F. nucleatum strains derived from saliva of CRC patients (Fig. S2) were analyzed. The top table shows the pattern of the CRISPR-associated regions of each strain. The numbers in the table are PCR product sizes shown in light (small sized) to darker (large sized) colors. In Patient D, five strains (A2, A3, A4, A5, and P3) were distinguished, although one strain ( V1 ) lacked the CRISPR-associated regions. In Patient E, four strains (A6, P4, P6, and P7) were distinguished, although two strains ( P5 and V2 ) lacked the CRISPR-associated regions. In Patient F, two strains ( P8 and P9 ) were distinguished, although one strain ( V3 ) lacked the CRISPR regions. In Patient G, one strain ( P10 ) was distinguished from each strain, although two strains ( V4 and V5 ) lacked the CRISPR regions. In Patient M, four strains (A12, A13, P19, and P20) were distinguished, although one strain ( V11 ) lacked the CRISPR regions.

Journal: Microbiology Spectrum

Article Title: Strain-level detection of Fusobacterium nucleatum in colorectal cancer specimens by targeting the CRISPR–Cas region

doi: 10.1128/spectrum.05123-22

Figure Lengend Snippet: Strain-level discrimination of F. nucleatum isolated from saliva of CRC patients by CRISPR-associated region-targeted PCR. Representative isolates of F. nucleatum strains derived from saliva of CRC patients (Fig. S2) were analyzed. The top table shows the pattern of the CRISPR-associated regions of each strain. The numbers in the table are PCR product sizes shown in light (small sized) to darker (large sized) colors. In Patient D, five strains (A2, A3, A4, A5, and P3) were distinguished, although one strain ( V1 ) lacked the CRISPR-associated regions. In Patient E, four strains (A6, P4, P6, and P7) were distinguished, although two strains ( P5 and V2 ) lacked the CRISPR-associated regions. In Patient F, two strains ( P8 and P9 ) were distinguished, although one strain ( V3 ) lacked the CRISPR regions. In Patient G, one strain ( P10 ) was distinguished from each strain, although two strains ( V4 and V5 ) lacked the CRISPR regions. In Patient M, four strains (A12, A13, P19, and P20) were distinguished, although one strain ( V11 ) lacked the CRISPR regions.

Techniques: Isolation, CRISPR, Derivative Assay

Detection of F. nucleatum strains from saliva of CRC patients by the F. nucleatum -strain genotyping PCR. ( a ) Detection of PCR amplicons from single and mixed multiple genomic DNA of F. nucleatum strains. From left to right, F. nucleatum subsp. animalis A3, A4, A5, A7 (Type I-B1); F. nucleatum subsp. polymorphum A-S11, P3, P7 (Type I-B2); F. nucleatum subsp. animalis A1, A8, F. nucleatum subsp. vincentii V7, F. nucleatum subsp. nucleatum N1 (Type II-A); F. nucleatum subsp. animalis A1, A3, F. nucleatum subsp. polymorphum P4 (Type III-A). Origins of many isolates are shown in Fig. S2. Strains A7, A8, and V7, A-S11, and A1, which are not in Fig. S2, were isolated from saliva of Patient H, saliva of Patient A, and CRC of Patient C, respectively, in our previous study . ( b ) Single genomic DNA of isolate or total genomic DNA extracted from saliva (the origin of the isolates) was used as a template (Patient M). The lane of total saliva (left, “S” in yellow) contains the bands identified in the isolates (blue arrowhead) and the amplicons derived from other non-isolates (pink arrowhead). Data are not available for strain V2 of Patient E and A12 of Patient M because these do not have all CRISPR types (see ).

Journal: Microbiology Spectrum

Article Title: Strain-level detection of Fusobacterium nucleatum in colorectal cancer specimens by targeting the CRISPR–Cas region

doi: 10.1128/spectrum.05123-22

Figure Lengend Snippet: Detection of F. nucleatum strains from saliva of CRC patients by the F. nucleatum -strain genotyping PCR. ( a ) Detection of PCR amplicons from single and mixed multiple genomic DNA of F. nucleatum strains. From left to right, F. nucleatum subsp. animalis A3, A4, A5, A7 (Type I-B1); F. nucleatum subsp. polymorphum A-S11, P3, P7 (Type I-B2); F. nucleatum subsp. animalis A1, A8, F. nucleatum subsp. vincentii V7, F. nucleatum subsp. nucleatum N1 (Type II-A); F. nucleatum subsp. animalis A1, A3, F. nucleatum subsp. polymorphum P4 (Type III-A). Origins of many isolates are shown in Fig. S2. Strains A7, A8, and V7, A-S11, and A1, which are not in Fig. S2, were isolated from saliva of Patient H, saliva of Patient A, and CRC of Patient C, respectively, in our previous study . ( b ) Single genomic DNA of isolate or total genomic DNA extracted from saliva (the origin of the isolates) was used as a template (Patient M). The lane of total saliva (left, “S” in yellow) contains the bands identified in the isolates (blue arrowhead) and the amplicons derived from other non-isolates (pink arrowhead). Data are not available for strain V2 of Patient E and A12 of Patient M because these do not have all CRISPR types (see ).

Techniques: Isolation, Derivative Assay, CRISPR

Application of the F. nucleatum -strain genotyping PCR to clinical trials. ( a ) Detection of identical strains of F. nucleatum from both CRC specimen and saliva obtained from patients with CRC by F. nucleatum -strain genotyping PCR. In two of the three patients (O, P, and Q), the presence of identical strain was confirmed by both the culture method and F. nucleatum -strain genotyping PCR. Left panel, detection of strain-specific DNA amplicons from genomic DNA of the CRC tissue and saliva obtained from a patient with CRC (Type I-B2). A same-size amplicon (red or blue arrowhead) was detected from both the CRC tissue and saliva. Two amplicons (yellow arrowhead), which were detected only in the saliva, were also CRISPR-associated regions derived from other Fusobacterium strains. Right panel, PCR amplicons from a single genomic DNA of isolates, which were identified as identical F. nucleatum strains, obtained from the same specimens. In Patient Q, no identical strains were detected from both CRC specimen and saliva samples. Lanes of isolates were cut and pasted from the original gel pictures (Fig. S13) for clarity. M, marker (DNA ladder: 0.1 kbp to 20 kbp). ( b ) Monitoring of the dynamics of F. nucleatum strains in saliva specimen from patients with colorectal adenoma and periodontitis, who were given oral care. Genomic DNAs derived from saliva at pre- and post-oral care were analyzed by the F. nucleatum -strain genotyping PCR (Type II-A). PCR amplicons derived from strain removed or decreased substantially by the oral care are shown as red arrowheads.

Journal: Microbiology Spectrum

Article Title: Strain-level detection of Fusobacterium nucleatum in colorectal cancer specimens by targeting the CRISPR–Cas region

doi: 10.1128/spectrum.05123-22

Figure Lengend Snippet: Application of the F. nucleatum -strain genotyping PCR to clinical trials. ( a ) Detection of identical strains of F. nucleatum from both CRC specimen and saliva obtained from patients with CRC by F. nucleatum -strain genotyping PCR. In two of the three patients (O, P, and Q), the presence of identical strain was confirmed by both the culture method and F. nucleatum -strain genotyping PCR. Left panel, detection of strain-specific DNA amplicons from genomic DNA of the CRC tissue and saliva obtained from a patient with CRC (Type I-B2). A same-size amplicon (red or blue arrowhead) was detected from both the CRC tissue and saliva. Two amplicons (yellow arrowhead), which were detected only in the saliva, were also CRISPR-associated regions derived from other Fusobacterium strains. Right panel, PCR amplicons from a single genomic DNA of isolates, which were identified as identical F. nucleatum strains, obtained from the same specimens. In Patient Q, no identical strains were detected from both CRC specimen and saliva samples. Lanes of isolates were cut and pasted from the original gel pictures (Fig. S13) for clarity. M, marker (DNA ladder: 0.1 kbp to 20 kbp). ( b ) Monitoring of the dynamics of F. nucleatum strains in saliva specimen from patients with colorectal adenoma and periodontitis, who were given oral care. Genomic DNAs derived from saliva at pre- and post-oral care were analyzed by the F. nucleatum -strain genotyping PCR (Type II-A). PCR amplicons derived from strain removed or decreased substantially by the oral care are shown as red arrowheads.

Techniques: Clinical Proteomics, Amplification, CRISPR, Derivative Assay, Marker

Overview of the CRISPR region-targeted PCR for the identification of F. nucleatum at the strain level and its application. ( a ) Sampling and preservation. Identification of bacteria at the strain level using cryopreserved samples without the need for conventional culture methods. ( b ) Procedure, interpretation, and application. Total DNA extracted from specimens and purified was analyzed by the F. nucleatum -strain genotyping PCR comprising the CRISPR-associated region targeting PCR, and the PCR using a primer set for nested PCR was performed followed by electrophoresis. An amplicon of the same size is found from both CRC and saliva, indicating that the identical strain is present in both CRC and saliva. Using this method, researchers can investigate the percentage of CRC promoted by oral F. nucleatum or prevent CRC recurrence. ( c ) For example, dynamic monitoring of the identical strain in both CRC and saliva, which is likely to be a pathogenic strain, during clinical follow-up. The effect of removal or reduction of this strain in oral cavity can then be investigated.

Journal: Microbiology Spectrum

Article Title: Strain-level detection of Fusobacterium nucleatum in colorectal cancer specimens by targeting the CRISPR–Cas region

doi: 10.1128/spectrum.05123-22

Figure Lengend Snippet: Overview of the CRISPR region-targeted PCR for the identification of F. nucleatum at the strain level and its application. ( a ) Sampling and preservation. Identification of bacteria at the strain level using cryopreserved samples without the need for conventional culture methods. ( b ) Procedure, interpretation, and application. Total DNA extracted from specimens and purified was analyzed by the F. nucleatum -strain genotyping PCR comprising the CRISPR-associated region targeting PCR, and the PCR using a primer set for nested PCR was performed followed by electrophoresis. An amplicon of the same size is found from both CRC and saliva, indicating that the identical strain is present in both CRC and saliva. Using this method, researchers can investigate the percentage of CRC promoted by oral F. nucleatum or prevent CRC recurrence. ( c ) For example, dynamic monitoring of the identical strain in both CRC and saliva, which is likely to be a pathogenic strain, during clinical follow-up. The effect of removal or reduction of this strain in oral cavity can then be investigated.

Techniques: CRISPR, Sampling, Preserving, Bacteria, Purification, Nested PCR, Electrophoresis, Amplification

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