Journal: Scientific Reports

Article Title: Bioelectrochemical conversion of CO 2 to value added product formate using engineered Methylobacterium extorquens

doi: 10.1038/s41598-018-23924-z

Figure Lengend Snippet: Diagram of the gene-knockout system for the construction of M. extorquens AM1 mutants. Sequences located on both sides of the target gene in the genome of M. extorquens AM1 were amplified by PCR and cloned into pCM184. The target gene was exchanged with the loxP-kan-loxP gene through homologous allelic recombination between M. extorquens AM1 and pCM184. For extraction of pCM184, M. extorquens AM1 mutants (Δtarget, KanR) were cultivated in media without kanamycin. Next, cre recombinase was expressed on pCM157 to eliminate the kanamycin resistance gene between loxP genes through site-specific recombination.

Article Snippet: When M. extorquens AM1 was transformed with pCM157 (Addgene plasmid 45863), cre recombinase expressed on pCM157 extracted the kanamycin gene between the loxP sites through site-specific recombination.

Techniques: Gene Knockout, Amplification, Clone Assay

Journal: Scientific Reports

Article Title: Bioelectrochemical conversion of CO 2 to value added product formate using engineered Methylobacterium extorquens

doi: 10.1038/s41598-018-23924-z

Figure Lengend Snippet: Bacterial strains and their plasmids for knockout or for recombinant expression.

Article Snippet: When M. extorquens AM1 was transformed with pCM157 (Addgene plasmid 45863), cre recombinase expressed on pCM157 extracted the kanamycin gene between the loxP sites through site-specific recombination.

Techniques: Knock-Out, Recombinant, Expressing, Plasmid Preparation

Journal: Journal of Clinical Medicine

Article Title: Rotator Cuff Tenocytes Differentiate into Hypertrophic Chondrocyte-Like Cells to Produce Calcium Deposits in an Alkaline Phosphatase-Dependent Manner

doi: 10.3390/jcm8101544

Figure Lengend Snippet: Assessment of mineral deposition induced by tenocyte-like cells (TLC) after 21 days in the osteogenic medium and characterization of mineralizing cells. ( A ) Assessment of mineral deposition by alizarin red S staining after 21 days ( N = 5). Quantification of the bound stain after solubilization with formic acid for the two conditions (reading at 450 nm) and representative images of the wells. CT: control medium; TLC: tenocyte-like cells; OM: osteogenic medium. Scale bar = 1 mm. ( B ) Gene expression by TLC cultured 21 days in the osteogenic medium or in the control medium (N = 3 to 5). 1: Study of TNAP (tissue non-specific alkaline phosphatase) and ENPP1 (ectonucleotide pyrophosphatase/phosphodiesterase 1) levels of expression. 2: Study of osteoblast markers: RUNX2, BGLAP (bone gamma-carboxyglutamate protein), BSP (Bone SialoProtein) and SPP1 (secreted phosphoProtein 1). 3: Study of chondrocyte markers: SOX9, COL2A1, ACAN (aggrecan), COMP (cartilage oligomeric matrix protein), COL10A1, and MMP13 (matrix metallopeptidase 13). 4: Study of tenocyte markers: SCX (scleraxis), MKX (Mohawk homeobox), TNMD (tenomodulin), COL1A1 and COL3A1. * = p < 0.05.

Article Snippet: Sections were incubated with primary antibodies targeting RUNX2 (Novus Biologicals NBP1-77461, 1/200), SOX9 (ab3697, 1/400, Abcam, UK), Type II collagen (COL2A1) (sc-52658, 1/1000, Santa Cruz Biotechnology, USA), Type X collagen (COL10) (ab49945, 1/1000, Abcam), tissue non-specific alkaline phosphatase (TNAP) (ab108337, 1/1000, Abcam), ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) (ab40003, 1/2000, Abcam), Mohawk homeobox (MKX) (NBP2-45863, 1/400, Novus Biologicals, USA), CD31 (ab28364, 1/100, Abcam), CD68 (M0876, 1/400, Dako, USA), and cleaved caspase-3 (9664, 1/400, Cell Signaling, Netherlands).

Techniques: Staining, Expressing, Cell Culture