Journal: Cell reports

Article Title: Modulation of Peptidoglycan Synthesis by Recycled Cell Wall Tetrapeptides

doi: 10.1016/j.celrep.2020.107578

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Penta-His antibody QIAGEN Cat#34660; RRID:AB_2619735 Anti-Mouse IgG Sigma Cat#A9044; RRID:AB_258431 Bacterial and Virus Strains All bacterial strains are listed in Table S1 This study Table S1 Chemicals, Peptides, and Recombinant Proteins D-Methionine Sigma Cat#M9375 L-Methionine Merck Cat#105707 Critical Commercial Assays Ni-NTA Agarose QIAGEN Cat#30210 Bio-Rad Protein Assay Dye Reagent Concentrate Bio-Rad Cat#500-0006 GeneJET Genomic DNA Purification Kit Thermo Fisher Scientific Cat#K0721 Qubit® dsDNA HS Assay Kit Thermo Fisher Scientific Cat# Q32851 Nextera XT DNA Library Preparation Kit Illumina Cat#FC-131-1024 MiSeq Reagent Kit v2 (50-cycles) Illumina Cat#MS-102-2001 Deposited Data Tn-seq experiments_Raw data This study https://www.ncbi.nlm.nih.gov BioProject ID: PRJNA623082 WGS suppressor mutants_Raw data This study https://www.ncbi.nlm.nih.gov BioProject ID: PRJNA623074 Oligonucleotides All oligonucleotides are detailed in Tables S2 – S4 This study Tables S2 – S4 Recombinant DNA pET28b:: vc2153 -6xHis This study FC281 pET28b:: ldcA -6xHis This study FC2025 pET22b:: vca0337 -6xHis This study FC2662 pET28b:: slt70 -6xHis ( E. coli ) Espaillat et al., 2016 FC1860 pET28b:: ldtA -6xHis Cava et al., 2011 FC1168 pET28b::AHA1477-6xHis This study FC2772 pET28b::PMI1557-6xHis This study FC2773 pET28b::STM1800-6xHis This study FC2774 pBAD 33 :: ldcA- 6xHis This study FC2753 pBAD 18-Km :: ldtA- 6xHis This study FC2759 pBAD 18-Km :: murA- 6xHis This study FC2838 pBAD 18-Km :: murB- 6xHis This study FC2839 pBAD 18-Km :: murC- 6xHis This study FC2840 pBAD 18-Km :: murE- 6xHis This study FC2841 pBAD 18-Km :: murF- 6xHis This study FC2842 Software and Algorithms GraphPad Prism GraphPad Software, Inc. https://www.graphpad.com Zeiss Zen Blue Software Zeiss https://www.zeiss.com/corporate/int/home.html ImageJ https://imagej.nih.gov/ij/ https://imagej.nih.gov/ij/ NCBI blast https://www.ncbi.nlm.nih.gov/BLAST/ https://www.ncbi.nlm.nih.gov/BLAST/ Galaxy server tools Afgan et al., 2016 https://usegalaxy.org/ Open in a separate window KEY RESOURCES TABLE.

Techniques: Virus, Recombinant, DNA Purification, DNA Library Preparation, Software

Journal: Applied and Environmental Microbiology

Article Title: Transposase-Mediated Chromosomal Integration of Exogenous Genes in Acidithiobacillus ferrooxidans

doi: 10.1128/AEM.01381-18

Figure Lengend Snippet: The transposon integration sites were confirmed by amplification of the region of the genome flanking the insertion sites. Panel A shows the genomic PCR amplification scheme for the AFKI1 strain. The primers flanking the insertion site were designed from the published genome sequence for ATCC 23270. In the wild-type genome, the PCR amplicon is of a known size. In the transposon-integrated strains, the same PCR primers were used to amplify the transposon region as well, resulting in a 3.5-kb increase in amplicon size. Panel B shows the results of the method applied to the three transposon locations identified in the mutant strains and visualized on an agarose gel, confirming the successful identification of integration sites.

Article Snippet: Using the NCBI Nucleotide blastx program, sequences were compared against the published ATCC 23270 genome to identify integration loci. table ft1 table-wrap mode="anchored" t5 TABLE 2 caption a7 Primer Sequence (5′–3′) Tn5Fwd TAT TAT CTG CGG CCG CCA TCG ACT GCA CGG TGC AC Tn5Rev AGA TAT CTG CGG CCG CTG TCA CTT T Tn5Rev2 AGA TAT CTC GCG GCC GCA AAA AGG CCA TCC GTC AGG ATG HypTnpFwd TAC ACA AGT AGC GTC GCA TGC CAT CGA CTG CAC HypTnpRev TTA GGC GGG CTA CTA TCT AGA TGT CAC TTT GCT TGA TAT ATG AGA ATT ATT TAA C pBAMFwd GAC GCT ACT TGT GTA CTG TCT CTT ATA CAC ATC TGA CGT CTT GTG T pBAMRev TAG TAG CCC GCC TAA TGA GCG pBAM2F AAG CGG GGT AAG CGC AAG AAT pBAM2R ATC GCC CAT GTT ATG CAG AAA tn1RFwd CCA CTA CCG GCA AGT TCT CCG tn2RFwd CAG TTC ACC GAC ACC AAA GGT G tn1RRev TAT GAA GAT GCA TGA GCC GGT C tn1LFwd TCG TCG ACC GAG CTT TTG C tn1LRev GAA AGA GGA TGC GCC GAA AGT G tn2LRev GGG AAA GCT CTT CGC CGA AC tnFSeq TGC ACA GCC ATA CCA CAG CTT C tnRSeq GGC TAC AGC TCG TTT CAC GCT G AFKI1Fwd TCG CCG TTC GTT TTC TCG AFKI1Rev GCC ACC GCA TCC AGT AAT C AFKI2Fwd ATG GTT CAC ACC GAA ATC AAT GC AFKI2Rev CAT CCA TGC TAC AGC CTA AGT TGC C AFKI3Fwd CCT GAT GTA GTC GTT GGC GTC C AFKI3Rev GTT CGT CAA CAG CAA AGT GGA AC Open in a separate window Primers used in this study (iii) Confirmation of integration loci.

Techniques: Amplification, Sequencing, Mutagenesis, Agarose Gel Electrophoresis

Journal: Applied and Environmental Microbiology

Article Title: Transposase-Mediated Chromosomal Integration of Exogenous Genes in Acidithiobacillus ferrooxidans

doi: 10.1128/AEM.01381-18

Figure Lengend Snippet: Location of the chromosomal integration sites. Panel A shows the integration loci for KDC-integrated strains. The insertion locus identifies the 9-bp sequence duplicated by the transposase to insert the transposon. Panel B shows the approximate locations of the transposon insertions for the mutant strains in relation to the whole A. ferrooxidans 23270 genome.

Article Snippet: Using the NCBI Nucleotide blastx program, sequences were compared against the published ATCC 23270 genome to identify integration loci. table ft1 table-wrap mode="anchored" t5 TABLE 2 caption a7 Primer Sequence (5′–3′) Tn5Fwd TAT TAT CTG CGG CCG CCA TCG ACT GCA CGG TGC AC Tn5Rev AGA TAT CTG CGG CCG CTG TCA CTT T Tn5Rev2 AGA TAT CTC GCG GCC GCA AAA AGG CCA TCC GTC AGG ATG HypTnpFwd TAC ACA AGT AGC GTC GCA TGC CAT CGA CTG CAC HypTnpRev TTA GGC GGG CTA CTA TCT AGA TGT CAC TTT GCT TGA TAT ATG AGA ATT ATT TAA C pBAMFwd GAC GCT ACT TGT GTA CTG TCT CTT ATA CAC ATC TGA CGT CTT GTG T pBAMRev TAG TAG CCC GCC TAA TGA GCG pBAM2F AAG CGG GGT AAG CGC AAG AAT pBAM2R ATC GCC CAT GTT ATG CAG AAA tn1RFwd CCA CTA CCG GCA AGT TCT CCG tn2RFwd CAG TTC ACC GAC ACC AAA GGT G tn1RRev TAT GAA GAT GCA TGA GCC GGT C tn1LFwd TCG TCG ACC GAG CTT TTG C tn1LRev GAA AGA GGA TGC GCC GAA AGT G tn2LRev GGG AAA GCT CTT CGC CGA AC tnFSeq TGC ACA GCC ATA CCA CAG CTT C tnRSeq GGC TAC AGC TCG TTT CAC GCT G AFKI1Fwd TCG CCG TTC GTT TTC TCG AFKI1Rev GCC ACC GCA TCC AGT AAT C AFKI2Fwd ATG GTT CAC ACC GAA ATC AAT GC AFKI2Rev CAT CCA TGC TAC AGC CTA AGT TGC C AFKI3Fwd CCT GAT GTA GTC GTT GGC GTC C AFKI3Rev GTT CGT CAA CAG CAA AGT GGA AC Open in a separate window Primers used in this study (iii) Confirmation of integration loci.

Techniques: Sequencing, Mutagenesis