Sequencing:Article Title: Corazonin links social role and metabolic-stress responses in honey bee workers
Article Snippet: .. Phusion High-Fidelity DNA polymerase (New England BioLabs) was used with the specific primers (Table S3) to produce the selected sequence in two different orientations with a 16-bp 5’ overhang needed for ligation using the In-Fusion HD Cloning system (Takara Bio): PCR products were purified using the NucleoSpin PCR purification kit, before being directionally inserted into the MCS of the pBluescript SK (+) vector downstream of the T7 promoter. .. Bacterial transformation was accomplished according to standard protocols, before using a QIAGEN Plasmid Midi Kit to isolate the plasmids from clonally selected cells.
Ligation:Article Title: Corazonin links social role and metabolic-stress responses in honey bee workers
Article Snippet: .. Phusion High-Fidelity DNA polymerase (New England BioLabs) was used with the specific primers (Table S3) to produce the selected sequence in two different orientations with a 16-bp 5’ overhang needed for ligation using the In-Fusion HD Cloning system (Takara Bio): PCR products were purified using the NucleoSpin PCR purification kit, before being directionally inserted into the MCS of the pBluescript SK (+) vector downstream of the T7 promoter. .. Bacterial transformation was accomplished according to standard protocols, before using a QIAGEN Plasmid Midi Kit to isolate the plasmids from clonally selected cells.
Cloning:Article Title: Corazonin links social role and metabolic-stress responses in honey bee workers
Article Snippet: .. Phusion High-Fidelity DNA polymerase (New England BioLabs) was used with the specific primers (Table S3) to produce the selected sequence in two different orientations with a 16-bp 5’ overhang needed for ligation using the In-Fusion HD Cloning system (Takara Bio): PCR products were purified using the NucleoSpin PCR purification kit, before being directionally inserted into the MCS of the pBluescript SK (+) vector downstream of the T7 promoter. .. Bacterial transformation was accomplished according to standard protocols, before using a QIAGEN Plasmid Midi Kit to isolate the plasmids from clonally selected cells.
Polymerase Chain Reaction:Article Title: Corazonin links social role and metabolic-stress responses in honey bee workers
Article Snippet: .. Phusion High-Fidelity DNA polymerase (New England BioLabs) was used with the specific primers (Table S3) to produce the selected sequence in two different orientations with a 16-bp 5’ overhang needed for ligation using the In-Fusion HD Cloning system (Takara Bio): PCR products were purified using the NucleoSpin PCR purification kit, before being directionally inserted into the MCS of the pBluescript SK (+) vector downstream of the T7 promoter. .. Bacterial transformation was accomplished according to standard protocols, before using a QIAGEN Plasmid Midi Kit to isolate the plasmids from clonally selected cells.
Article Title: A multifunctional PKS-NRPS megasynthetase in nematodes produces divergent signaling molecules that control different developmental arrests
Article Snippet: .. Construction and imaging of a Ppa-pks-1p::turboRFP reporter strain in P. pacificus To make the Ppa-pks-1p::turboRFP transcriptional reporter plasmid, 3 kb of the Ppa-pks-1 promoter was PCR-amplified from a synthetic gene fragment (Twist Bioscience) using Phusion High-Fidelity DNA Polymerase (New England Biolabs 0530) and inserted into the PZH009 (Ppa-egl-20p::turboRFP) plasmid40 using Gibson Assembly (New England Biolabs 2611). ..
Article Title: Optimized cloning and expression of recombinant human type I collagen in mammalian cells using Ubiquitous Chromatin Opening Elements for enhanced protein production
Article Snippet: .. High-fidelity PCR amplification of the COL1A1 gene was performed using Phusion High-Fidelity DNA Polymerase (Cat. #M0530S, New England Biolabs, USA). .. Restriction enzyme digestion employed FseI (Cat. #R0588S, New England Biolabs, USA) and NheI (Cat. #R0131S, New England Biolabs, USA).
Article Title: Glutamine sequestration promotes tumor growth and immune evasion through elevation of arginine methylation in hepatocellular carcinoma
Article Snippet: .. The libraries were amplified with 15 cycles of PCR using Phusion High-Fidelity DNA Polymerase (New England Biolabs) and index primers. .. Final library concentration and size distribution were quantified using a Qubit 4.0 Fluorometer (Thermo Fisher Scientific) and an Agilent 5300 Bioanalyzer.
Article Title: P450 cyptide synthase KwwB catalyzes non-native cross-link at Tyr-C3-Trp-N1 and tolerates leader peptide mutations.
Article Snippet: .. These plasmids containing mutated residue were amplified with Phusion high-fidelity DNA polymerase (New England Biolabs) in PCR thermal cycler (Eppendorf), followed by digestion with enzyme DpnI (New England Biolabs), gel purification with DNA extraction kit (Bio Basic), DNA 5’ phosphorylation with T4 polynucleotide kinase (New England Biolabs), and DNA ligation with T4 DNA ligase (New England Biolabs). .. The cloned plasmids were transformed into E. coli DH5α, then harvested with Monarch plasmid miniprep kit (New England Biolabs), and sequence verified at Genewiz from Azenta Life Sciences.
Article Title: ATM promotes nucleolytic processing of reversed replication forks during DNA interstrand cross-link repair
Article Snippet: .. Recovered DNA was amplified by PCR with the primers RA302 and RA303 (see Supplementary Table 1) and Phusion High-Fidelity DNA Polymerase (NEB #M0530). .. Amplicons were submitted to Azenta for paired-end NGS (Illumina MiSeq, 2 × 250bp).
Article Title: Molecular Characterization of Soritesidine Reveals a Novel Marine Sponge DNase I-like Toxin
Article Snippet: The concentration and purity of the isolated nucleic acids were determined using a NanoDrop spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA), and RNA integrity was assessed by electrophoresis on a 0.8% agarose gel. .. Unless otherwise stated, all PCR amplifications were performed by nested PCR using Phusion High-Fidelity DNA Polymerase (New England Biolabs, Ipswich, MA, USA). .. PCR products were purified from agarose gels using the AxyPrep DNA Gel Extraction Kit (Axygen, Union City, CA, USA) and subjected to DNA sequencing.
Purification:Article Title: Corazonin links social role and metabolic-stress responses in honey bee workers
Article Snippet: .. Phusion High-Fidelity DNA polymerase (New England BioLabs) was used with the specific primers (Table S3) to produce the selected sequence in two different orientations with a 16-bp 5’ overhang needed for ligation using the In-Fusion HD Cloning system (Takara Bio): PCR products were purified using the NucleoSpin PCR purification kit, before being directionally inserted into the MCS of the pBluescript SK (+) vector downstream of the T7 promoter. .. Bacterial transformation was accomplished according to standard protocols, before using a QIAGEN Plasmid Midi Kit to isolate the plasmids from clonally selected cells.
Plasmid Preparation:Article Title: Corazonin links social role and metabolic-stress responses in honey bee workers
Article Snippet: .. Phusion High-Fidelity DNA polymerase (New England BioLabs) was used with the specific primers (Table S3) to produce the selected sequence in two different orientations with a 16-bp 5’ overhang needed for ligation using the In-Fusion HD Cloning system (Takara Bio): PCR products were purified using the NucleoSpin PCR purification kit, before being directionally inserted into the MCS of the pBluescript SK (+) vector downstream of the T7 promoter. .. Bacterial transformation was accomplished according to standard protocols, before using a QIAGEN Plasmid Midi Kit to isolate the plasmids from clonally selected cells.
Article Title: A multifunctional PKS-NRPS megasynthetase in nematodes produces divergent signaling molecules that control different developmental arrests
Article Snippet: .. Construction and imaging of a Ppa-pks-1p::turboRFP reporter strain in P. pacificus To make the Ppa-pks-1p::turboRFP transcriptional reporter plasmid, 3 kb of the Ppa-pks-1 promoter was PCR-amplified from a synthetic gene fragment (Twist Bioscience) using Phusion High-Fidelity DNA Polymerase (New England Biolabs 0530) and inserted into the PZH009 (Ppa-egl-20p::turboRFP) plasmid40 using Gibson Assembly (New England Biolabs 2611). ..
Imaging:Article Title: A multifunctional PKS-NRPS megasynthetase in nematodes produces divergent signaling molecules that control different developmental arrests
Article Snippet: .. Construction and imaging of a Ppa-pks-1p::turboRFP reporter strain in P. pacificus To make the Ppa-pks-1p::turboRFP transcriptional reporter plasmid, 3 kb of the Ppa-pks-1 promoter was PCR-amplified from a synthetic gene fragment (Twist Bioscience) using Phusion High-Fidelity DNA Polymerase (New England Biolabs 0530) and inserted into the PZH009 (Ppa-egl-20p::turboRFP) plasmid40 using Gibson Assembly (New England Biolabs 2611). ..
Amplification:
Article Title: Optimized cloning and expression of recombinant human type I collagen in mammalian cells using Ubiquitous Chromatin Opening Elements for enhanced protein production
Article Snippet: .. High-fidelity PCR amplification of the COL1A1 gene was performed using Phusion High-Fidelity DNA Polymerase (Cat. #M0530S, New England Biolabs, USA). .. Restriction enzyme digestion employed FseI (Cat. #R0588S, New England Biolabs, USA) and NheI (Cat. #R0131S, New England Biolabs, USA).
Article Title: Glutamine sequestration promotes tumor growth and immune evasion through elevation of arginine methylation in hepatocellular carcinoma
Article Snippet: .. The libraries were amplified with 15 cycles of PCR using Phusion High-Fidelity DNA Polymerase (New England Biolabs) and index primers. .. Final library concentration and size distribution were quantified using a Qubit 4.0 Fluorometer (Thermo Fisher Scientific) and an Agilent 5300 Bioanalyzer.
Article Title: P450 cyptide synthase KwwB catalyzes non-native cross-link at Tyr-C3-Trp-N1 and tolerates leader peptide mutations.
Article Snippet: .. These plasmids containing mutated residue were amplified with Phusion high-fidelity DNA polymerase (New England Biolabs) in PCR thermal cycler (Eppendorf), followed by digestion with enzyme DpnI (New England Biolabs), gel purification with DNA extraction kit (Bio Basic), DNA 5’ phosphorylation with T4 polynucleotide kinase (New England Biolabs), and DNA ligation with T4 DNA ligase (New England Biolabs). .. The cloned plasmids were transformed into E. coli DH5α, then harvested with Monarch plasmid miniprep kit (New England Biolabs), and sequence verified at Genewiz from Azenta Life Sciences.
Article Title: ATM promotes nucleolytic processing of reversed replication forks during DNA interstrand cross-link repair
Article Snippet: .. Recovered DNA was amplified by PCR with the primers RA302 and RA303 (see Supplementary Table 1) and Phusion High-Fidelity DNA Polymerase (NEB #M0530). .. Amplicons were submitted to Azenta for paired-end NGS (Illumina MiSeq, 2 × 250bp).
Residue:Article Title: P450 cyptide synthase KwwB catalyzes non-native cross-link at Tyr-C3-Trp-N1 and tolerates leader peptide mutations.
Article Snippet: .. These plasmids containing mutated residue were amplified with Phusion high-fidelity DNA polymerase (New England Biolabs) in PCR thermal cycler (Eppendorf), followed by digestion with enzyme DpnI (New England Biolabs), gel purification with DNA extraction kit (Bio Basic), DNA 5’ phosphorylation with T4 polynucleotide kinase (New England Biolabs), and DNA ligation with T4 DNA ligase (New England Biolabs). .. The cloned plasmids were transformed into E. coli DH5α, then harvested with Monarch plasmid miniprep kit (New England Biolabs), and sequence verified at Genewiz from Azenta Life Sciences.
Gel Purification:Article Title: P450 cyptide synthase KwwB catalyzes non-native cross-link at Tyr-C3-Trp-N1 and tolerates leader peptide mutations.
Article Snippet: .. These plasmids containing mutated residue were amplified with Phusion high-fidelity DNA polymerase (New England Biolabs) in PCR thermal cycler (Eppendorf), followed by digestion with enzyme DpnI (New England Biolabs), gel purification with DNA extraction kit (Bio Basic), DNA 5’ phosphorylation with T4 polynucleotide kinase (New England Biolabs), and DNA ligation with T4 DNA ligase (New England Biolabs). .. The cloned plasmids were transformed into E. coli DH5α, then harvested with Monarch plasmid miniprep kit (New England Biolabs), and sequence verified at Genewiz from Azenta Life Sciences.
DNA Extraction:Article Title: P450 cyptide synthase KwwB catalyzes non-native cross-link at Tyr-C3-Trp-N1 and tolerates leader peptide mutations.
Article Snippet: .. These plasmids containing mutated residue were amplified with Phusion high-fidelity DNA polymerase (New England Biolabs) in PCR thermal cycler (Eppendorf), followed by digestion with enzyme DpnI (New England Biolabs), gel purification with DNA extraction kit (Bio Basic), DNA 5’ phosphorylation with T4 polynucleotide kinase (New England Biolabs), and DNA ligation with T4 DNA ligase (New England Biolabs). .. The cloned plasmids were transformed into E. coli DH5α, then harvested with Monarch plasmid miniprep kit (New England Biolabs), and sequence verified at Genewiz from Azenta Life Sciences.
Phospho-proteomics:Article Title: P450 cyptide synthase KwwB catalyzes non-native cross-link at Tyr-C3-Trp-N1 and tolerates leader peptide mutations.
Article Snippet: .. These plasmids containing mutated residue were amplified with Phusion high-fidelity DNA polymerase (New England Biolabs) in PCR thermal cycler (Eppendorf), followed by digestion with enzyme DpnI (New England Biolabs), gel purification with DNA extraction kit (Bio Basic), DNA 5’ phosphorylation with T4 polynucleotide kinase (New England Biolabs), and DNA ligation with T4 DNA ligase (New England Biolabs). .. The cloned plasmids were transformed into E. coli DH5α, then harvested with Monarch plasmid miniprep kit (New England Biolabs), and sequence verified at Genewiz from Azenta Life Sciences.
DNA Ligation:Article Title: P450 cyptide synthase KwwB catalyzes non-native cross-link at Tyr-C3-Trp-N1 and tolerates leader peptide mutations.
Article Snippet: .. These plasmids containing mutated residue were amplified with Phusion high-fidelity DNA polymerase (New England Biolabs) in PCR thermal cycler (Eppendorf), followed by digestion with enzyme DpnI (New England Biolabs), gel purification with DNA extraction kit (Bio Basic), DNA 5’ phosphorylation with T4 polynucleotide kinase (New England Biolabs), and DNA ligation with T4 DNA ligase (New England Biolabs). .. The cloned plasmids were transformed into E. coli DH5α, then harvested with Monarch plasmid miniprep kit (New England Biolabs), and sequence verified at Genewiz from Azenta Life Sciences.
Nested PCR:Article Title: Molecular Characterization of Soritesidine Reveals a Novel Marine Sponge DNase I-like Toxin
Article Snippet: The concentration and purity of the isolated nucleic acids were determined using a NanoDrop spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA), and RNA integrity was assessed by electrophoresis on a 0.8% agarose gel. .. Unless otherwise stated, all PCR amplifications were performed by nested PCR using Phusion High-Fidelity DNA Polymerase (New England Biolabs, Ipswich, MA, USA). .. PCR products were purified from agarose gels using the AxyPrep DNA Gel Extraction Kit (Axygen, Union City, CA, USA) and subjected to DNA sequencing.
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