Journal: Frontiers in Microbiology

Article Title: A Bacterial Expression Vector Archive (BEVA) for Flexible Modular Assembly of Golden Gate-Compatible Vectors

doi: 10.3389/fmicb.2018.03345

Figure Lengend Snippet: Bacterial Expression Vector Archive (BEVA) system for the modular assembly of bacterial vectors. (A) Vector modules are shown in the order they are assembled in a vector construction reaction. Their position in the final construct is labeled. Sequences (5′→ 3′) used to anneal the fragments in order are shown (color-coded) at junctions between modules and remain in the final construct as a scar. (B) Modules at each position. Position 1: cloning sites – Golden Gate level 1 and Golden Gate level 2; position 2: antibiotic resistance – neomycin-, gentamicin- and tetracycline-resistance cassettes; position 3: origins of replication and transfer – RK2 with oriT and pBBR1 with oriT ; positions 4–6: accessory modules – par locus for stable plasmid maintenance in the absence of antibiotic selection. Endlinker modules were designed to circularize the plasmid.

Article Snippet: The BEVA vectors for use in Golden Gate cloning (level 1: pOGG005, pOGG024 and pOGG026, and level 2: pOGG216) have been made available through Addgene (Table ).

Techniques: Expressing, Plasmid Preparation, Construct, Labeling, Cloning, Selection

Journal: Frontiers in Microbiology

Article Title: A Bacterial Expression Vector Archive (BEVA) for Flexible Modular Assembly of Golden Gate-Compatible Vectors

doi: 10.3389/fmicb.2018.03345

Figure Lengend Snippet: (A) Schematic map of pOGG024 level 1 broad-host range, medium copy number vector constructed with BEVA for free-living gene expression in laboratory. (B) Schematic map of plasmid pOPS0359 containing a cloned gene sfGFP under the control of a taurine-inducible promoter from Sinorhizobium meliloti in pOGG024. (C–F) Evaluation of pOPS0359 performance as robust free-living gene expression compared to the functionally analogous pLMB509 plasmid. (C,D) Fluorescence detection from GFP expression and (E,F) fitness performance by measurement of OD 595 when grown without (0 mM) or with (0.5, 1.0, 10, or 40 mM) taurine. For all graphs, error bars are standard error of the mean (SEM).

Article Snippet: The BEVA vectors for use in Golden Gate cloning (level 1: pOGG005, pOGG024 and pOGG026, and level 2: pOGG216) have been made available through Addgene (Table ).

Techniques: Plasmid Preparation, Construct, Gene Expression, Clone Assay, Control, Fluorescence, Expressing

Journal: Frontiers in Microbiology

Article Title: A Bacterial Expression Vector Archive (BEVA) for Flexible Modular Assembly of Golden Gate-Compatible Vectors

doi: 10.3389/fmicb.2018.03345

Figure Lengend Snippet: (A) Schematic map of pOGG026 level 1 broad-host range, lower copy number vector constructed with BEVA for stable environmental gene expression in the absence of antibiotic selection. (B,C) Schematic map of plasmids containing a cloned (B) gusA (pOPS0253) and (C) celB (pOPS0254) under the control of nifH gene promoter (P nifH ). (D) Pea nodules formed by Rlv3841[pOPS253] stained with Magenta-glucA. Marker gene gusA is just expressed in nodules. (E) Pea nodules formed by Rlv3841[pOPS254] and (F) bean nodules formed by CIAT899[pOPS254] stained with X-gal after thermal treatment. Marker genes gusA and celB are just expressed in nodules.

Article Snippet: The BEVA vectors for use in Golden Gate cloning (level 1: pOGG005, pOGG024 and pOGG026, and level 2: pOGG216) have been made available through Addgene (Table ).

Techniques: Plasmid Preparation, Construct, Gene Expression, Selection, Clone Assay, Control, Staining, Marker

Journal: Frontiers in Microbiology

Article Title: A Bacterial Expression Vector Archive (BEVA) for Flexible Modular Assembly of Golden Gate-Compatible Vectors

doi: 10.3389/fmicb.2018.03345

Figure Lengend Snippet: (A) Schematic map of pOGG216 level 2 broad-host range, medium copy number vector constructed with BEVA for multi-gene expression. (B) Schematic map of dual reporter plasmid pOPS0754 which encodes IPTG-inducible sfGFP cloned in Forward Position 1 and constitutively mCherry cloned in Forward Position 2. (C,E,G) Green fluorescence detection (scale, 5,000–31,000). (D,F) Red fluorescence detection (scale, 3,000–29,000 cps). (C,D) Rlv3841 without fluorescence. Rlv3841[pOPS0754] expressing IPTG-inducible sfGFP grown in media (E) containing 0.5 mM IPTG or (G) without IPTG. (F) Rlv3841[pOPS0754] with constitutively expressed mCherry.

Article Snippet: The BEVA vectors for use in Golden Gate cloning (level 1: pOGG005, pOGG024 and pOGG026, and level 2: pOGG216) have been made available through Addgene (Table ).

Techniques: Plasmid Preparation, Construct, Gene Expression, Clone Assay, Fluorescence, Expressing