Journal:

Article Title: Biosynthesis of Lipoteichoic Acid in Lactobacillus rhamnosus : Role of DltD in d -Alanylation

doi:

Figure Lengend Snippet: Bacterial strains, phages, and plasmids used in this study a

Article Snippet: All bacterial strains, phages, and plasmids used in this study are listed in Table . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Strain, plasmid, or phage Description Source or reference Strains L. rhamnosus 7469 Formerly named L. casei ATCC 7469 ATCC L. casei 102S L. casei ATCC 393 cured from plasmids B. Chassy L. casei 102S dltD :: erm Em r ; dltD interrupted by erm This study E. coli XL-1 Blue MRF′ Δ( mcrA ) 183 Δ ( mcrCB-hsdSMR-mrr ) 173end1A supE44thi-1 recA1 gyrA96 relA1lac [F′ proAB lac q Z ΔM15 Tn 10 (Tet r )] Stratagene E. coli BL21(DE3) pLysS F − ompT hsdS B (r B − m B − ) gal dcm (DE3)pLysS Novagene E. coli DH5α F − f80d lacZ ΔM15 Δ( lacZYA-argF ) U169 deoR recA1 endA1 hsdR17 (r k − m k + ) supE44 λ − thi-1 girA96 relA1 Gibco-BRL Phages (phagemids) λZAPII [Bluescript SK( − )] Ap r cloning vector Stratagene VCSM13 Helper phage Stratagene λZAPII(−)DE43 4.3 kb of L. rhamnosus DNA containing dltA , dltB , dltC , and partial dltD 10 λZAPII(−)A65 6.5 kb of L. rhamnosus DNA containing partial dltB , dltC , dltD , and downstream flanking region This study Plasmids pET-3d Expression vector Novagene pDltD Expression plasmid; dltD cloned into pET-3d This study pΔDltD Expression plasmid; Δ dltD cloned into pET-3d This study pVE6006 Em r Cm r Ts shuttle vector 30 pNZ123 Cm r shuttle vector 46 pΔDltD/ erm 1.2-kb Eco ICR erm fragment of pVE6006 joined to Bam HI blunt-ended pΔDltD in direct orientation to Δ dltD This study pNZ123/ dlt 4.895-kb fragment containing dlt operon cloned into Hin dIII of pNZ123 This study Open in a separate window a Em r , erythromycin resistant; Cm r , choramphenicol resistant; Ap r , ampicillin resistant; ATCC, American Type Culture Collection.

Techniques: Plasmid Preparation, Cloning, Expressing, Clone Assay

Journal:

Article Title: Biosynthesis of Lipoteichoic Acid in Lactobacillus rhamnosus : Role of DltD in d -Alanylation

doi:

Figure Lengend Snippet: Incorporation of d -[ 14 C]alanine into permeabilized cells and membranes from L. casei 102S ( dltD + ) and its dltD :: erm mutant

Article Snippet: All bacterial strains, phages, and plasmids used in this study are listed in Table . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Strain, plasmid, or phage Description Source or reference Strains L. rhamnosus 7469 Formerly named L. casei ATCC 7469 ATCC L. casei 102S L. casei ATCC 393 cured from plasmids B. Chassy L. casei 102S dltD :: erm Em r ; dltD interrupted by erm This study E. coli XL-1 Blue MRF′ Δ( mcrA ) 183 Δ ( mcrCB-hsdSMR-mrr ) 173end1A supE44thi-1 recA1 gyrA96 relA1lac [F′ proAB lac q Z ΔM15 Tn 10 (Tet r )] Stratagene E. coli BL21(DE3) pLysS F − ompT hsdS B (r B − m B − ) gal dcm (DE3)pLysS Novagene E. coli DH5α F − f80d lacZ ΔM15 Δ( lacZYA-argF ) U169 deoR recA1 endA1 hsdR17 (r k − m k + ) supE44 λ − thi-1 girA96 relA1 Gibco-BRL Phages (phagemids) λZAPII [Bluescript SK( − )] Ap r cloning vector Stratagene VCSM13 Helper phage Stratagene λZAPII(−)DE43 4.3 kb of L. rhamnosus DNA containing dltA , dltB , dltC , and partial dltD 10 λZAPII(−)A65 6.5 kb of L. rhamnosus DNA containing partial dltB , dltC , dltD , and downstream flanking region This study Plasmids pET-3d Expression vector Novagene pDltD Expression plasmid; dltD cloned into pET-3d This study pΔDltD Expression plasmid; Δ dltD cloned into pET-3d This study pVE6006 Em r Cm r Ts shuttle vector 30 pNZ123 Cm r shuttle vector 46 pΔDltD/ erm 1.2-kb Eco ICR erm fragment of pVE6006 joined to Bam HI blunt-ended pΔDltD in direct orientation to Δ dltD This study pNZ123/ dlt 4.895-kb fragment containing dlt operon cloned into Hin dIII of pNZ123 This study Open in a separate window a Em r , erythromycin resistant; Cm r , choramphenicol resistant; Ap r , ampicillin resistant; ATCC, American Type Culture Collection.

Techniques:

Journal:

Article Title: Biosynthesis of Lipoteichoic Acid in Lactobacillus rhamnosus : Role of DltD in d -Alanylation

doi:

Figure Lengend Snippet: Formation of d-alanyl–Dcp and d-alanyl–ACP in the presence of DltD+, DltD−, and DltD−/DltD+ membranes. Nondenaturing PAGE (15%) was used to monitor the amount of either d-[14C]alanyl–Dcp or d-[14C]alanyl–ACP according to the method of Heaton and Neuhaus (19). The reactions were performed in 250-μl mixtures containing 8 U of Dcl, 5 nmol of either Dcp or ACP, 5 mM MgCl2, 5 mM DTT, 30 mM bis-Tris buffer (pH 6.5), 10 mM ATP, and 110 μM d[14C]alanine (46 mCi/mmol) in the absence of membranes (w/o) and in the presence of membranes (130 μg) from L. casei 102S (dltD+), L. casei 102S (dltD::erm), and the dltD mutant which had been complemented with pNZ123/dlt (dltD::erm/dltD+). All reaction mixtures were incubated for 1 h at 37°C.

Article Snippet: All bacterial strains, phages, and plasmids used in this study are listed in Table . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Strain, plasmid, or phage Description Source or reference Strains L. rhamnosus 7469 Formerly named L. casei ATCC 7469 ATCC L. casei 102S L. casei ATCC 393 cured from plasmids B. Chassy L. casei 102S dltD :: erm Em r ; dltD interrupted by erm This study E. coli XL-1 Blue MRF′ Δ( mcrA ) 183 Δ ( mcrCB-hsdSMR-mrr ) 173end1A supE44thi-1 recA1 gyrA96 relA1lac [F′ proAB lac q Z ΔM15 Tn 10 (Tet r )] Stratagene E. coli BL21(DE3) pLysS F − ompT hsdS B (r B − m B − ) gal dcm (DE3)pLysS Novagene E. coli DH5α F − f80d lacZ ΔM15 Δ( lacZYA-argF ) U169 deoR recA1 endA1 hsdR17 (r k − m k + ) supE44 λ − thi-1 girA96 relA1 Gibco-BRL Phages (phagemids) λZAPII [Bluescript SK( − )] Ap r cloning vector Stratagene VCSM13 Helper phage Stratagene λZAPII(−)DE43 4.3 kb of L. rhamnosus DNA containing dltA , dltB , dltC , and partial dltD 10 λZAPII(−)A65 6.5 kb of L. rhamnosus DNA containing partial dltB , dltC , dltD , and downstream flanking region This study Plasmids pET-3d Expression vector Novagene pDltD Expression plasmid; dltD cloned into pET-3d This study pΔDltD Expression plasmid; Δ dltD cloned into pET-3d This study pVE6006 Em r Cm r Ts shuttle vector 30 pNZ123 Cm r shuttle vector 46 pΔDltD/ erm 1.2-kb Eco ICR erm fragment of pVE6006 joined to Bam HI blunt-ended pΔDltD in direct orientation to Δ dltD This study pNZ123/ dlt 4.895-kb fragment containing dlt operon cloned into Hin dIII of pNZ123 This study Open in a separate window a Em r , erythromycin resistant; Cm r , choramphenicol resistant; Ap r , ampicillin resistant; ATCC, American Type Culture Collection.

Techniques: Mutagenesis, Incubation

Journal: BMC Cancer

Article Title: Lactobacillus casei combined with Lactobacillus reuteri alleviate pancreatic cancer by inhibiting TLR4 to promote macrophage M1 polarization and regulate gut microbial homeostasis

doi: 10.1186/s12885-023-11557-z

Figure Lengend Snippet: L. casei & L. reuteri inhibits the proliferation, migration, invasion of pancreatic cancer cells and pancreatic cancer cell-induced M2 polarization of macrophages. ( A ) MTT assay of MIA PaCa-2, Panc-1, BxPC-3, and AsPC-1 cells. ( B ) RT-qPCR analysis of TLR4 and MyD88 expressions in BxPC-3 cells. ( C ) CCK-8 assay of BxPC-3 cells. ( D ) Transwell migration and invasion assay of BxPC-3 cells. ( E ) Representative chart of flow cytometric analysis of CD86 + or CD206 + macrophages. ( F ) Statistical chart of flow cytometric analysis of CD86 + or CD206 + macrophages. ( G ) WB analysis of Arg-1 and iNOS in macrophages. Lactobacillus represents cells treated with L. casei & L. reuteri. * p < 0.05

Article Snippet: L. casei (ATCC 39392) and L. reuteri (ATCC 23272) were purchased from the American Type Culture Collection.

Techniques: Migration, MTT Assay, Quantitative RT-PCR, CCK-8 Assay, Invasion Assay

Journal: BMC Cancer

Article Title: Lactobacillus casei combined with Lactobacillus reuteri alleviate pancreatic cancer by inhibiting TLR4 to promote macrophage M1 polarization and regulate gut microbial homeostasis

doi: 10.1186/s12885-023-11557-z

Figure Lengend Snippet: L. casei & L. reuteri influences pancreatic cancer cells and macrophage polarization by regulating TLR4. ( A ) RT-qPCR analysis of TLR4 and MyD88 expressions in BxPC-3 cells. ( B ) Statistical chart of flow cytometric analysis of CD86 + or CD206 + macrophages. ( C ) Representative chart of flow cytometric analysis of CD86 + or CD206 + macrophages. ( D ) WB analysis of Arg-1 and iNOS in macrophages. ( E ) CCK-8 assay of BxPC-3 cells. ( F ) Transwell migration and invasion assay of BxPC-3 cells. * p < 0.05 versus control group, # p < 0.05 versus LPS group

Article Snippet: L. casei (ATCC 39392) and L. reuteri (ATCC 23272) were purchased from the American Type Culture Collection.

Techniques: Quantitative RT-PCR, CCK-8 Assay, Migration, Invasion Assay, Control

Journal: BMC Cancer

Article Title: Lactobacillus casei combined with Lactobacillus reuteri alleviate pancreatic cancer by inhibiting TLR4 to promote macrophage M1 polarization and regulate gut microbial homeostasis

doi: 10.1186/s12885-023-11557-z

Figure Lengend Snippet: L. casei & L. reuteri represses pancreatic cancer growth and promotes M1 macrophage polarization by inhibiting TLR4. ( A ) Nude mice were xenografted with BxPC-3 cells (PC group) and treated with L. casei & L. reuteri (Lactobacillus group) or TAK-242 (TAK-242 group). The tumor sizes are shown. ( B ) The tumor volumes (left) and tumor weights (right) of nude mice. ( C ) Body weights of nude mice. ( D ) RT-qPCR analysis of TLR4 and MyD88 expressions in tumor tissues. ( E ) IHC staining for Ki67 in tumor tissues. Scale bar: 100 μm (upper), 25 μm (lower). ( F ) IF staining for CD86 and CD206 in tumor tissues. ( G ) IF staining for Arg-1 and iNOS in tumor tissues. ( H ) Flow cytometric analysis of CD86 + or CD206 + macrophages in tumor tissues. * p < 0.05 versus PC group

Article Snippet: L. casei (ATCC 39392) and L. reuteri (ATCC 23272) were purchased from the American Type Culture Collection.

Techniques: Quantitative RT-PCR, Immunohistochemistry, Staining

Journal: BMC Cancer

Article Title: Lactobacillus casei combined with Lactobacillus reuteri alleviate pancreatic cancer by inhibiting TLR4 to promote macrophage M1 polarization and regulate gut microbial homeostasis

doi: 10.1186/s12885-023-11557-z

Figure Lengend Snippet: L. casei & L. reuteri regulates gut microbial homeostasis by inhibiting TLR4. ( A ) ASV analysis of the relative abundance of species. ( B ) Alpha diversity of gut microbiota. ( C ) PCoA analysis of gut microbiota. ( D ) Beta diversity of gut microbiota. ( E ) The relative abundance of gut microbiota at phylum (left) and genus (right) level. ( F ) Differences of relative abundance of specific gut microbiota at genus level. * p < 0.05 versus control group, # p < 0.05 versus PC group

Article Snippet: L. casei (ATCC 39392) and L. reuteri (ATCC 23272) were purchased from the American Type Culture Collection.

Techniques: Control

Journal: BMC Cancer

Article Title: Lactobacillus casei combined with Lactobacillus reuteri alleviate pancreatic cancer by inhibiting TLR4 to promote macrophage M1 polarization and regulate gut microbial homeostasis

doi: 10.1186/s12885-023-11557-z

Figure Lengend Snippet: L. casei & L. reuteri regulates gut microbial homeostasis by inhibiting TLR4. ( A ) PCA scores plot for metabolic profiles. ( B ) PLS-DA plot for metabolic profiles. ( C ) The heat map of the metabolic features

Article Snippet: L. casei (ATCC 39392) and L. reuteri (ATCC 23272) were purchased from the American Type Culture Collection.

Techniques: