Photorhabdus Asymbiotica Subsp, supplied by ATCC, used in various techniques. Bioz Stars score: 93/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "Photorhabdus Virulence Cassettes: extracellular multi-protein needle complexes for delivery of small protein effectors into host cells"
Article Title: Photorhabdus Virulence Cassettes: extracellular multi-protein needle complexes for delivery of small protein effectors into host cells
Figure Legend Snippet: ( A ) Boxplots of the mean GC content across 16 different pvc operons of Photorhabdus . The GC was calculated for each of the 16 structural loci (clustered by annotated/predicted function and syntenic position in operon consistent with the nomenclature devised in this paper). GC content itself was calculated via a bespoke script, outputting data to be visualised in RStudio. Data was plotted and the step-function fit (black dashed line) was calculated using the mean GC value for each locus via the cumSeg package for breakpoint estimation in genomic sequences. Diamonds represent mean locus GC. Beige box shows the source genome mean (dotted line) GC content and standard deviation (upper and lower box bounds). Grey box shows the operon GC mean (dotted line) and standard deviation (upper and lower box bounds). ( B ) Box plots of amino acid similarity across homologous protein sequences for these same 16 operons. Amino acid sequences were clustered together as in (A), by annotation and syntenic position. Global Multiple Sequence Alignments were created with CLUSTAL Omega, using the default parameters (Gonnet transition matrix, gap open penalty 6 bits, gap extend 1 bit). Pairwise amino acid alignment scores were extracted from the CLUSTALO output and plotted in RStudio via bespoke scripts. Diamonds indicate mean pairwise alignment scores. Dots indicate pairwise values that are outliers, beyond 1.5 X the interquartile range (as automatically calculated by the ggplot2 package). ( C ) A map of the model class I Pa ATCC43949 PVC pnf operon showing two effector genes in the payload region in red/orange.
Techniques Used: Genomic Sequencing, Standard Deviation, Sequencing
Figure Legend Snippet: A representative selection of images for 4 time points, for P. asymbiotica PB68.1 ( Pa PB68 ) harbouring (A) the Pa PB68 PVC pnf pvc1 promoter fusion construct or (B) pAGAG negative control reporter plasmid with no promoter. For (A) quadruplicate images are displayed vertically as representative of the whole slide sample. Key to qualitative fluorescence indication: “-” is no fluorescence, “++” is low level fluorescence in many cells or a few brighter cells, “+++” is intermediate to high fluorescence in almost all cells, or some very bright isolated cells.
Techniques Used: Selection, Construct, Negative Control, Plasmid Preparation, Fluorescence, Isolation
Figure Legend Snippet: (A) RT-PCR analysis of gene transcription of various genes of the Pa ATCC43949 PVC pnf operon over time in vitro at insect (28°C) and human (37°C) relevant temperatures and in vivo during Manduca sexta ( Ms ) infection. Lane key; lanes 1, 2 and 3 (blue) represent in vitro growth in aerated LB at 28°C for 4, 8 and 24h respectively; lanes 4, 5 and 6 (red) are growth in aerated LB at 37°C for 4, 8 and 24h; lane 7 (black) is growth in LB at 28°C for 16h; lanes 8 and 9 (green) are from 3h and 6h post infection blood of Ms infected with P. asymbiotica at 28°C. (B) Map of the operon showing RT-PCR target genes in black. The lane-colour coded bars above the ORFs summarise in which conditions gene transcription could be detected. Note pvc8 and pvc14 mRNA could not be detected from infected Ms and the pnf mRNA was only detected after 6h of infection. (C) RT-PCR signals for pvc15 and pnf from infected insects with the rpsM (ribosomal subunit protein S13) loading control. Lanes represent (in order); 4h growth in LB at 28°C; 4h growth in Grace’s insect medium supplemented with 20% (v/v) Ms hemolymph; 3h and 6h post infection ex vivo blood of Ms infected with P. asymbiotica at 28°C.
Techniques Used: Reverse Transcription Polymerase Chain Reaction, In Vitro, In Vivo, Infection, Ex Vivo
Figure Legend Snippet: PVC operons in relation to putative regions of horizontal gene transfer as identified by Alien Hunter. ( A ) The PVC pnf and ( B ) the PVC cif operons are highlighted by the blue rectangle. Alien Hunter regions of HGT are designated by the features in tones of red. In red are the regions with the highest score and thus probability for HGT whilst in white are the regions with the lowest scores. (C ) The P. asymbiotica ATCC43949 chromosome. The first concentric circle denotes genes on the forward strand while the second circle denotes genes in the reverse strand. In dark green are the PVC operons. The third circle shows regions of HGT as identified by Alien Hunter.
Figure Legend Snippet: ( A ) Similarity between PVCs and two diverse protein secretion systems, (i) the P. luteoviolacea mac gene cluster and (ii) The type-VI secretion system (T6SS) from Burkholderia mallei. Homologous protein sequences are coloured coded. (B) Three classes of PVC structural operons observed in the genomes of Photorhabdus and members of other genera. Types 1-3 are exemplified by PVC pnf , PVC lopT and PVC PaTox respectively. Homologous genes are colour coded. Red arrows represent variations relative to the representative type I PVC pnf operon of P. asymbiotica ATCC43949, pvc1 (PAU_03353) to pvc16 (PAU_03338). Predicted functions of individual Pvc proteins based on homology to known proteins can be seen in FigS1C . The boxed “GGCGGTAGNNT” or “CAGGTTGXTGCGGTAGCTAT” sequences represent positions of the conserved RfaH anti-termination protein and cryptic operator sequences respectively. Square brackets above certain genes indicate apparent translational coupling. More specifically; *1 indicates coupling in PVC pnf and PVC cif of Pl TT01 , Pa ATCC43949 , Pa PB68 and Pa Kingscliff and in the Serratia entomophila afp operon in addition to an uncharacterised PVC in Yersinia ruckeri ATCC 29473. **1 indicates these genes are not coupled in Pa Ki ngscliff . *2 indicates coupling in PVC lopT of Pl TT01 , Pa ATCC43949 , Pa PB68 and Pa Kingscliff . **2 indicates these genes are not coupled in Pa Kingscliff . *3 indicates coupling in PVC Patox of Pa ATCC43949 and Pa Kingscliff (although pvc11 possibly contains a frame-shift in Pa Kingscliff ). The pvc3 is also deleted in Pa Kingscliff .