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Characterization of sporulation dynamics of Pseudoclostridium thermosuccinogenes using flow cytometry

Single-cell analysis of microbial population heterogeneity is a fast growing research area in microbiology due to its potential to identify and quantify the impact of subpopulations on microbial performance in, for example, industrial biotechnology, environmental biology, and pathogenesis. Although several tools have been developed, determination of population heterogenity in anaerobic bacteria, especially spore-forming clostridia species has been amply studied.

hAlbumin FRET 50
AH1005-50 1 Kit
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hInsulin FRET 50
IH1017-50 1 Kit
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hInsulin TRF 50
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mIgG TRF 50
MIGTR1020-50 1 Kit
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rIgG FRET 50
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TruStrip Sample Transfer Strips, 50-ul, 50/Pk (without sample tracking dye)
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BNUM0943-50 50uL
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Rv2450c (RpfE)-50
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Rv3875 (esxA)-50
AR02-P0008-50 50ug
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Rv2031c(HspX)-50
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Rv3131 (NfnB)-50
AR02-P0010-50 50ug
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ERLENMEYER FLASK, 50 ML, PP
4985P-50 6/pk
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ERLENMEYER FLASK, 50 ML, PMP
4990P-50 6/pk
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Description: Reusable Plastics; Reusable Erl Flasks

Rv2769c (PE27 protein)-50
AR02-P0001-50 50ug
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TRF-1-1-1003A 1 Kit
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slingshot homolog 1 (SSH1)(MBP-his tagged)-50
AR09-P0023-50 50ug
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DNA Gel Extraction Kit - 50 Columns
31030-50 1kit
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Cytokeratin 5/8(C-50) Antibody
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TRF-1-1-1001A 1 Kit
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A118-0.5MG 0.5 mg
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M-phase inducer phosphatase 1 (CDC25A)(MBP-his tagged)-50
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Rv0350 (Heat Shock Protein 70)-50
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protein tyrosine phosphatase, non-receptor type 1 (PTPN1)(His tagged)-50
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Antigen Retrieval Solution, Citrate pH 6.0 (50 ml, 100X)
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PTPRF(His tagged) / LAR(His tagged)-50
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ExoQuick-CG cGMP-grade Exosome Isolation reagent (50 ml)
EXOCG50A-1 50 ml
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  • Category: Exosomes

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5-00619 4 x 1mg Ask for price

Adrenomedullin (1-50), rat
HY-P1534 1mg
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Antigen Retrieval Solution, Tris-EDTA pH 9.0 (50 ml, 100X)
ARS9-50 5 L
EUR 286.00

BasicExoRNA? Basic RNA extraction kit (Pre-isolated exosomes, 50 reactions)
K1222-50
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Adrenomedullin (Adrenomedullin (ADM/AM, 11-50) (rat)
SP-100047-1 1 mg
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slingshot homolog 3 (SSH3)(MBP-his tagged)-50
AR09-P0024-50 50ug
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AXYPREP MAG DYECLEAN KIT- 50 ML - 5000 PREPS
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Human IgG/RF-adsorbent (Adsorb 50 serum samples of 100 ul)
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PTPRC(His tagged) / Human CD45 (LCA)(His tagged)-50
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GP3911-1G 1 g
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Description: Bioscience Mag Beads; Magnetic Fragment Select

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Description: Reservoirs; Reservoirs - Axygen

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dual specificity phosphatase-like 15 (DUSP15)(MBP-his tagged)-50
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dual specificity phosphatase-like 18 (DUSP18)(MBP-his tagged)-50
AR09-P0019-50 50ug
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hSKRP1 mRNA for SKRP1 (NE1, DUSP19)(MBP-his tagged)-50
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dual specificity phosphatase-like 21 (DUSP21)(MBP-his tagged)-50
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dual specificity phosphatase-like 26 (DUSP26)(MBP-his tagged)-50
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protein tyrosine phosphatase zeta-polypeptide (PTPRZ)(MBP-his tagged)-50
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C2000-50 500/pack
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100 bp DNA Ladder Plus, ready-to-use, 50 ug, 0.5 ug per lane
Z6030001-1 100 lanes
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50 L RPMI 1640, POWDER WITH L-GLUTAMINE, WITHOUT SODIUM BICARBONATE
50-020-PC 50 L/pk
EUR 92.00
Description: Media Catalog; Classical Media

In this study we applied single cell analysis techniques such as flow cytometry (FCM) and fluorescence-assisted cell sorting (FACS) on the spore-forming succinate producer Pseudoclostridium thermosuccinogenes. By combining FCM and FACS with fluorescent staining, we differentiated and enriched all sporulation-related morphologies of P. thermosuccinogenes.

To evaluate the presence of metabolically active vegetative cells, a blend of the dyes propidium iodide (PI) and carboxy fluorescein diacetate (cFDA) tested best. Side scatter (SSC-H) in combination with metabolic indicator cFDA dye provided the best separation of sporulation populations.

Based on this protocol, we successfully determined culture heterogeneity of P. thermosuccinogenes by discriminating between mature spores, forespores, dark and bright phase endospores, and vegetative cells populations. Henceforth, this methodology can be applied to further study sporulation dynamics and its impact on fermentation performance and product formation by P. thermosuccinogenes.

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