Journal:

Article Title: Association of Actinobacillus pleuropneumoniae Capsular Polysaccharide with Virulence in Pigs

doi: 10.1128/IAI.71.6.3320-3328.2003

Figure Lengend Snippet: Bacterial strains and plasmids used

Article Snippet: Kanamycin and streptomycin were used at 85 and 80 μg/ml, respectively, for selection of A. pleuropneumoniae recombinant strains. table ft1 table-wrap mode="anchored" t5 TABLE 1. caption a7 Strain or plasmid Relevant genotype(s) or characteristic(s) Source or reference A. pleuropneumoniae strains 4074 Serotype 1 (ATCC 27088) ATCC a J45-100 Nonencapsulated mutant of serotype 5a 55 4074Δ cps 1N Nonencapsulated, nontypeable mutant of 4074; cps1AB Kan r This work 4074Δ cps 1B Nonencapsulated, nontypeable mutant of 4074; cps1B Kan r This work 4074Δ cps 1N(pAB cps 101) 4074Δ cps 1N containing pAB cps 101; serotype 1 + Str r This work 4074Δ cps 1N(pAB cps 55) 4074Δ cps 1N containing pAB cps 55; serotype 1 + Str r This work 4074Δ cps 1N(pJML cps 53) 4074Δ cps 1N containing pJML cps 53; serotype 1 + Str r This work E. coli XL1-Blue recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac ( F + proABlacI q ZΔM15 Tn 10 ); host for recombinant plasmids Stratagene Plasmids pBluescript II SK(+/−) 2.96-kb cloning vector; Amp r Stratagene pGSAp11 4.8-kb EcoR I fragment of 4074 cloned into pBluescript This work pMLAp53 8-kb EcoR I fragment of J45 cloned into pGEM-3z 55 pUC4K E. coli cloning vector; Amp r Kan r Pharmacia pAp12KB pGSAp11 with 0.7-kb Bgl II fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pGSAp11K pGSAp11 with 0.5-kb Nde I- SnaB I fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pLS88 Broad-host-range shuttle vector from H. ducreyii ; Str r Kan r 58 pAB cps 101 pLS88 containing cps1ABC in the Kan r site; Str r This work pAB cps 55 pLS88 containing cps5ABCDE in the Kan r site; Str r This work pJML cps 53 pLS88 containing cps5ABC in the Kan r site; Str r 55 Open in a separate window a American Type Culture Collection, Rockville, Md.

Techniques: Plasmid Preparation, Mutagenesis, Recombinant, Clone Assay

Journal:

Article Title: Association of Actinobacillus pleuropneumoniae Capsular Polysaccharide with Virulence in Pigs

doi: 10.1128/IAI.71.6.3320-3328.2003

Figure Lengend Snippet: Deletion of cps1AB in strain 4074Δcps1N. (A) The primers CPS1U2 and CPS1L3 were used to amplify the region deleted by mutagenesis of cps1AB. (B) The primers CPS1U3 and CPS1L4 were used to amplify the region not deleted by mutagenesis. The strains used were the serotype 5a knockout mutant strain J45-100 as a negative control (lanes 2), the serotype 1 parent strain 4074 (lanes 3), and the serotype 1 knockout mutant strain 4074Δcps1N (lanes 4).

Article Snippet: Kanamycin and streptomycin were used at 85 and 80 μg/ml, respectively, for selection of A. pleuropneumoniae recombinant strains. table ft1 table-wrap mode="anchored" t5 TABLE 1. caption a7 Strain or plasmid Relevant genotype(s) or characteristic(s) Source or reference A. pleuropneumoniae strains 4074 Serotype 1 (ATCC 27088) ATCC a J45-100 Nonencapsulated mutant of serotype 5a 55 4074Δ cps 1N Nonencapsulated, nontypeable mutant of 4074; cps1AB Kan r This work 4074Δ cps 1B Nonencapsulated, nontypeable mutant of 4074; cps1B Kan r This work 4074Δ cps 1N(pAB cps 101) 4074Δ cps 1N containing pAB cps 101; serotype 1 + Str r This work 4074Δ cps 1N(pAB cps 55) 4074Δ cps 1N containing pAB cps 55; serotype 1 + Str r This work 4074Δ cps 1N(pJML cps 53) 4074Δ cps 1N containing pJML cps 53; serotype 1 + Str r This work E. coli XL1-Blue recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac ( F + proABlacI q ZΔM15 Tn 10 ); host for recombinant plasmids Stratagene Plasmids pBluescript II SK(+/−) 2.96-kb cloning vector; Amp r Stratagene pGSAp11 4.8-kb EcoR I fragment of 4074 cloned into pBluescript This work pMLAp53 8-kb EcoR I fragment of J45 cloned into pGEM-3z 55 pUC4K E. coli cloning vector; Amp r Kan r Pharmacia pAp12KB pGSAp11 with 0.7-kb Bgl II fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pGSAp11K pGSAp11 with 0.5-kb Nde I- SnaB I fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pLS88 Broad-host-range shuttle vector from H. ducreyii ; Str r Kan r 58 pAB cps 101 pLS88 containing cps1ABC in the Kan r site; Str r This work pAB cps 55 pLS88 containing cps5ABCDE in the Kan r site; Str r This work pJML cps 53 pLS88 containing cps5ABC in the Kan r site; Str r 55 Open in a separate window a American Type Culture Collection, Rockville, Md.

Techniques: Mutagenesis, Knock-Out, Negative Control

Journal:

Article Title: Association of Actinobacillus pleuropneumoniae Capsular Polysaccharide with Virulence in Pigs

doi: 10.1128/IAI.71.6.3320-3328.2003

Figure Lengend Snippet: Qualitative determination of CP production. Latex beads conjugated to IgG specific for serotype 1 CP was used. The parent strain, 4074, producing serotype 1 CP, agglutinated with the beads, while the nonencapsulated mutant 4074Δcps1N did not.

Article Snippet: Kanamycin and streptomycin were used at 85 and 80 μg/ml, respectively, for selection of A. pleuropneumoniae recombinant strains. table ft1 table-wrap mode="anchored" t5 TABLE 1. caption a7 Strain or plasmid Relevant genotype(s) or characteristic(s) Source or reference A. pleuropneumoniae strains 4074 Serotype 1 (ATCC 27088) ATCC a J45-100 Nonencapsulated mutant of serotype 5a 55 4074Δ cps 1N Nonencapsulated, nontypeable mutant of 4074; cps1AB Kan r This work 4074Δ cps 1B Nonencapsulated, nontypeable mutant of 4074; cps1B Kan r This work 4074Δ cps 1N(pAB cps 101) 4074Δ cps 1N containing pAB cps 101; serotype 1 + Str r This work 4074Δ cps 1N(pAB cps 55) 4074Δ cps 1N containing pAB cps 55; serotype 1 + Str r This work 4074Δ cps 1N(pJML cps 53) 4074Δ cps 1N containing pJML cps 53; serotype 1 + Str r This work E. coli XL1-Blue recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac ( F + proABlacI q ZΔM15 Tn 10 ); host for recombinant plasmids Stratagene Plasmids pBluescript II SK(+/−) 2.96-kb cloning vector; Amp r Stratagene pGSAp11 4.8-kb EcoR I fragment of 4074 cloned into pBluescript This work pMLAp53 8-kb EcoR I fragment of J45 cloned into pGEM-3z 55 pUC4K E. coli cloning vector; Amp r Kan r Pharmacia pAp12KB pGSAp11 with 0.7-kb Bgl II fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pGSAp11K pGSAp11 with 0.5-kb Nde I- SnaB I fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pLS88 Broad-host-range shuttle vector from H. ducreyii ; Str r Kan r 58 pAB cps 101 pLS88 containing cps1ABC in the Kan r site; Str r This work pAB cps 55 pLS88 containing cps5ABCDE in the Kan r site; Str r This work pJML cps 53 pLS88 containing cps5ABC in the Kan r site; Str r 55 Open in a separate window a American Type Culture Collection, Rockville, Md.

Techniques: Mutagenesis

Journal:

Article Title: Association of Actinobacillus pleuropneumoniae Capsular Polysaccharide with Virulence in Pigs

doi: 10.1128/IAI.71.6.3320-3328.2003

Figure Lengend Snippet: Capsule production and degree of virulence in pigs of Ap serotype 1 isogenic strains

Article Snippet: Kanamycin and streptomycin were used at 85 and 80 μg/ml, respectively, for selection of A. pleuropneumoniae recombinant strains. table ft1 table-wrap mode="anchored" t5 TABLE 1. caption a7 Strain or plasmid Relevant genotype(s) or characteristic(s) Source or reference A. pleuropneumoniae strains 4074 Serotype 1 (ATCC 27088) ATCC a J45-100 Nonencapsulated mutant of serotype 5a 55 4074Δ cps 1N Nonencapsulated, nontypeable mutant of 4074; cps1AB Kan r This work 4074Δ cps 1B Nonencapsulated, nontypeable mutant of 4074; cps1B Kan r This work 4074Δ cps 1N(pAB cps 101) 4074Δ cps 1N containing pAB cps 101; serotype 1 + Str r This work 4074Δ cps 1N(pAB cps 55) 4074Δ cps 1N containing pAB cps 55; serotype 1 + Str r This work 4074Δ cps 1N(pJML cps 53) 4074Δ cps 1N containing pJML cps 53; serotype 1 + Str r This work E. coli XL1-Blue recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac ( F + proABlacI q ZΔM15 Tn 10 ); host for recombinant plasmids Stratagene Plasmids pBluescript II SK(+/−) 2.96-kb cloning vector; Amp r Stratagene pGSAp11 4.8-kb EcoR I fragment of 4074 cloned into pBluescript This work pMLAp53 8-kb EcoR I fragment of J45 cloned into pGEM-3z 55 pUC4K E. coli cloning vector; Amp r Kan r Pharmacia pAp12KB pGSAp11 with 0.7-kb Bgl II fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pGSAp11K pGSAp11 with 0.5-kb Nde I- SnaB I fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pLS88 Broad-host-range shuttle vector from H. ducreyii ; Str r Kan r 58 pAB cps 101 pLS88 containing cps1ABC in the Kan r site; Str r This work pAB cps 55 pLS88 containing cps5ABCDE in the Kan r site; Str r This work pJML cps 53 pLS88 containing cps5ABC in the Kan r site; Str r 55 Open in a separate window a American Type Culture Collection, Rockville, Md.

Techniques: Produced

Journal:

Article Title: Association of Actinobacillus pleuropneumoniae Capsular Polysaccharide with Virulence in Pigs

doi: 10.1128/IAI.71.6.3320-3328.2003

Figure Lengend Snippet: Bactericidal activity of precolostral calf serum for strains 4074, 4074Δcps1N, and J45-100. The percent viability of each strain was evaluated 0 and 60 min after incubation at 37°C with 25% precolostral calf serum. Each data point represents the mean for three separate experiments performed in duplicate.

Article Snippet: Kanamycin and streptomycin were used at 85 and 80 μg/ml, respectively, for selection of A. pleuropneumoniae recombinant strains. table ft1 table-wrap mode="anchored" t5 TABLE 1. caption a7 Strain or plasmid Relevant genotype(s) or characteristic(s) Source or reference A. pleuropneumoniae strains 4074 Serotype 1 (ATCC 27088) ATCC a J45-100 Nonencapsulated mutant of serotype 5a 55 4074Δ cps 1N Nonencapsulated, nontypeable mutant of 4074; cps1AB Kan r This work 4074Δ cps 1B Nonencapsulated, nontypeable mutant of 4074; cps1B Kan r This work 4074Δ cps 1N(pAB cps 101) 4074Δ cps 1N containing pAB cps 101; serotype 1 + Str r This work 4074Δ cps 1N(pAB cps 55) 4074Δ cps 1N containing pAB cps 55; serotype 1 + Str r This work 4074Δ cps 1N(pJML cps 53) 4074Δ cps 1N containing pJML cps 53; serotype 1 + Str r This work E. coli XL1-Blue recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac ( F + proABlacI q ZΔM15 Tn 10 ); host for recombinant plasmids Stratagene Plasmids pBluescript II SK(+/−) 2.96-kb cloning vector; Amp r Stratagene pGSAp11 4.8-kb EcoR I fragment of 4074 cloned into pBluescript This work pMLAp53 8-kb EcoR I fragment of J45 cloned into pGEM-3z 55 pUC4K E. coli cloning vector; Amp r Kan r Pharmacia pAp12KB pGSAp11 with 0.7-kb Bgl II fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pGSAp11K pGSAp11 with 0.5-kb Nde I- SnaB I fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pLS88 Broad-host-range shuttle vector from H. ducreyii ; Str r Kan r 58 pAB cps 101 pLS88 containing cps1ABC in the Kan r site; Str r This work pAB cps 55 pLS88 containing cps5ABCDE in the Kan r site; Str r This work pJML cps 53 pLS88 containing cps5ABC in the Kan r site; Str r 55 Open in a separate window a American Type Culture Collection, Rockville, Md.

Techniques: Activity Assay, Incubation

Journal:

Article Title: Association of Actinobacillus pleuropneumoniae Capsular Polysaccharide with Virulence in Pigs

doi: 10.1128/IAI.71.6.3320-3328.2003

Figure Lengend Snippet: Qualitative determination of CP production by latex beads conjugated to IgG specific for serotype 5 CP. Strains 4074 and 4074Δcps1N did not produce serotype 5 CP and did not agglutinate the beads, while the serotype 5 CP-producing chimeric strains 4074Δcps1N(pABcps55) and 4074Δcps1N(pJMLcps53) did agglutinate.

Article Snippet: Kanamycin and streptomycin were used at 85 and 80 μg/ml, respectively, for selection of A. pleuropneumoniae recombinant strains. table ft1 table-wrap mode="anchored" t5 TABLE 1. caption a7 Strain or plasmid Relevant genotype(s) or characteristic(s) Source or reference A. pleuropneumoniae strains 4074 Serotype 1 (ATCC 27088) ATCC a J45-100 Nonencapsulated mutant of serotype 5a 55 4074Δ cps 1N Nonencapsulated, nontypeable mutant of 4074; cps1AB Kan r This work 4074Δ cps 1B Nonencapsulated, nontypeable mutant of 4074; cps1B Kan r This work 4074Δ cps 1N(pAB cps 101) 4074Δ cps 1N containing pAB cps 101; serotype 1 + Str r This work 4074Δ cps 1N(pAB cps 55) 4074Δ cps 1N containing pAB cps 55; serotype 1 + Str r This work 4074Δ cps 1N(pJML cps 53) 4074Δ cps 1N containing pJML cps 53; serotype 1 + Str r This work E. coli XL1-Blue recA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac ( F + proABlacI q ZΔM15 Tn 10 ); host for recombinant plasmids Stratagene Plasmids pBluescript II SK(+/−) 2.96-kb cloning vector; Amp r Stratagene pGSAp11 4.8-kb EcoR I fragment of 4074 cloned into pBluescript This work pMLAp53 8-kb EcoR I fragment of J45 cloned into pGEM-3z 55 pUC4K E. coli cloning vector; Amp r Kan r Pharmacia pAp12KB pGSAp11 with 0.7-kb Bgl II fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pGSAp11K pGSAp11 with 0.5-kb Nde I- SnaB I fragment deleted and blunt ended and 1.2-kb Sal I-cut and blunt-ended Kan r cassette from pUC4K ligated This work pLS88 Broad-host-range shuttle vector from H. ducreyii ; Str r Kan r 58 pAB cps 101 pLS88 containing cps1ABC in the Kan r site; Str r This work pAB cps 55 pLS88 containing cps5ABCDE in the Kan r site; Str r This work pJML cps 53 pLS88 containing cps5ABC in the Kan r site; Str r 55 Open in a separate window a American Type Culture Collection, Rockville, Md.

Techniques:

Journal: Scientific Reports

Article Title: Mechanisms and consequences of casein kinase II and ankyrin-3 regulation of the epithelial Na + channel

doi: 10.1038/s41598-021-94118-3

Figure Lengend Snippet: Over-expression of CKII is sufficient to increase ENaC levels in the plasma membrane and activity. ( A ) Fluorescence micrographs of COS-7 cells expressing eYFP-ENaC in the absence (top) and presence of over-expression of CKII (bottom) prior to photobleaching (left) and 10 s (middle) and 10 min (right) after photobleaching. Cells were transfected with eYFP-ENaC and CKII plasmids, CK2alpha and CK2beta. Images were collected with TIRF microscopy. ( B ) Time course of relative FRAP at the plasma membrane for cells expressing eYFP-ENaC in the absence (black circles) and presence of over-expression of CKII (white triangles). Summary data from experiments (n = 6–9 cells from 2–3 distinct transfections) identical to those shown in ( A ). ( C ) Summary graph of relative FRAP 10 min after photobleaching in cells expressing eYFP-ENaC in the absence (black circles, gray bar) and presence (white triangles, white bar) of over-expression of CKII. Summary data from experiments identical to those shown in ( A ). *P < 0.05 vs. the absence of CKII over-expression. ( D ) Overlays of typical macroscopic current traces from representative CHO cells expressing mENaC in the absence (top) and presence of over-expression of CKII (bottom) before and after 10 µM amiloride (dotted lines). Currents elicited by voltage ramps stepped from a holding potential of 40 mV to 60 mV and ramped to − 100 mV. ( E ) Summary graph of ENaC activity (amiloride-sensitive current density at -100 mV) quantified in whole-cell voltage clamped CHO cells transfected with mENaC in the absence (black circles, gray bar) and presence (white triangles, white bar) of over-expression of CKII. Summary data from experiments (n = 9–19 cells from 3–5 distinct transfections) identical to those shown in ( D ). *P < 0.05 vs. the absence of CKII over-expression.

Article Snippet: The plasmids encoding pZW6 (CK2alpha) and pZW12 (CK2beta) were from Addgene (Addgene_27086 and Addgene_27088, respectively; Watertown, MA, USA).

Techniques: Over Expression, Clinical Proteomics, Membrane, Activity Assay, Fluorescence, Expressing, Transfection, Microscopy