Construction of Chlamydia Fluorescent Reporters

All constructs were created in the p2TK2SW2 plasmid background (38 (link)). Promoters and the ftsI ORF were amplified from C. trachomatis L2 (LGV Bu434) genomic DNA using the indicated primers (see Table S1 in the supplemental material). Fluorescent reporters were ordered as gBlocks and cloned using the In-Fusion HD EcoDry Cloning kit (TaKaRa). Promoter reporter constructs were created as previously described (10 (link), 39 (link)). The p2TK2SW2-E-ftsI3XFLAG was generated by replacing the Clover gene with the ftsI ORF in the previously created p2TK2SW2-E-Clover-3XFLAG plasmid (12 (link)). Dual promoter reporter cassettes [euoprom-mNG(LVA)_hctBprom-mKate2 and hctAprom-mEos3.2_hctBprom-mKate2] were then inserted upstream of E-ftsI-3XFLAG to produce the p2TK2SW2-E-ftsI-3XFLAG_euoprom-mNG(LVA)_hctBprom-mKate2, and p2TK2SW2-E-ftsI-3XFLAG_hctAprom-mEos3.2_hctBprom-mKate2 constructs.

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Chiarelli T.J., Grieshaber N.A., Appa C, & Grieshaber S.S. (2023). Computational Modeling of the Chlamydial Developmental Cycle Reveals a Potential Role for Asymmetric Division. mSystems, 8(2), e00053-23.

Publication 2023

In-Fusion HD EcoDry Cloning kit

Clover Gene Genomic Plasmid Primers

Corresponding Organization : University of Idaho

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Variable analysis

independent variables

Promoters and the ftsI ORF amplified from C. trachomatis L2 (LGV Bu434) genomic DNA using indicated primers

dependent variables

Expression of fluorescent reporters (mNG(LVA), mKate2, mEos3.2)

control variables

All constructs were created in the p2TK2SW2 plasmid background

controls

No positive or negative controls were explicitly mentioned.

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