pcDNA3.1-msEGFP1.8
(Plasmid #208619)

• Purpose: Mammalian expression of a EGFP variant. Mutations were adopted from (Addgene#135301, PMC7508230), named msEGFP1.8 because N-ter and C-ter features of that in msGFP2 (PMC7508230) not included.
• Depositing Lab: Quanfu Ma
• Publication: RBM24 interacts with RPL3 and regulates RPL3-eIF6 interaction (unpublished)

Backbone

• Vector backbone: pcDNA3.1 (-)
• Backbone manufacturer: Thermo Fisher (Invitrogen)
• Modifications to backbone: introduced some restriction site from pEGFP-C1 (between NheI and XhoI)
• Vector type: Mammalian Expression
• Selectable markers: Neomycin (select with G418)

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: msEGFP1.8
• Alt name: EGFP
• Alt name: GFP
• Alt name: monomer superfold enhanced green fluoresence protein
• Species: Synthetic; Aequorea victoria
• Mutation: (compared with pEGFP-C1, protein level) S30R, Y39N, F99T, N105T,M153T, V163A, I171V, A206K
• GenBank ID: U55763 AAB02576
• Promoter: CMV

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: NheI (not destroyed)
• 3′ cloning site: XhoI (not destroyed)
• 5′ sequencing primer: CMV-F
• 3′ sequencing primer: BGH-R

Resource Information

• Supplemental Documents:
  • pcDNA3.1-msEGFP1.8.gb
• A portion of this plasmid was derived from a plasmid made by: EGFP sequence clone from comercial plasmid pEGFP-C1 (TAKARA-Clontech), sites between NheI and XhoI in pEGFP-C1 were introduced along with EGFP. mutations were adopted from Benjamin Glick's work (Addgene#135301, PMC7508230), named msEGFP1.8 because N-ter and C-ter features of that in Benjamin Glick's work not included, thus 0.2 less than that in Benjamin Glick's work.

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

mutations were introduced as discribed in "https://www.neb.com/-/media/nebus/files/application-notes/appnote_improved_methods_for_sdm_using_nebuilder_hifi_dna_assembly_master_mix.pdf"
>mutagenesis_primers
EGFP-S30R+Y39N-F: 5' GTTCAGCGTGCGCGGCGAGGGCGAGGGCGATGCCACCAACGGCAAGC 3'
EGFP-S30R+Y39N-R: 5' GCTTGCCGTTGGTGGCATCGCCCTCGCCCTCGCCGCGCACGCTGAAC 3'
EGFP-F99T+N105T-F: 5' CACCATCACCTTCAAGGACGACGGCACCTACAAGACC 3'
EGFP-F99T+N105T-R: 5' GGTCTTGTAGGTGCCGTCGTCCTTGAAGGTGATGGTG 3'
EGFP-M153T-R: 5' TCTTCTGCTTGTCGGCGGTGATATAGACGTTG 3'
EGFP-M153T+V163A.o1: 5' CGTCTATATCACCGCCGACAAGCAGAAGAACGGCATCAAGGCCAACTTCAAGATC 3'
EGFP-V163A+I171V.o2: 5' CTGCCGTCCTCCACGTTGTGGCGGATCTTGAAGTTGGCCTTGATGCCGTTC 3'
EGFP-I171V-F: 5' AGATCCGCCACAACGTGGAGGACGGCAGCG 3'
EGFP-A206K-F: 5' CCTGAGCACCCAGTCCAAGCTGAGCAAAGACCCCA 3'
EGFP-A206K-R: 5' TGGGGTCTTTGCTCAGCTTGGACTGGGTGCTCAGG 3'