LOCUS       sequence-474162-        4245 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mGreenLantern expression to serve as a transfection marker, 
            EF1a-driven.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4245)
  AUTHORS   Love KS, Johnstone CP, Peterman EL, Gaglione S, Birnbaum ME, 
            Galloway KE
  TITLE     Model-guided design of microRNA-based gene circuits supports precise
            dosage of transgenic cargoes into diverse primary cells.
  JOURNAL   Cell Syst. 2025 Apr 22:101269. doi: 10.1016/j.cels.2025.101269.
  PUBMED    40300600
REFERENCE   2  (bases 1 to 4245)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Syst. 
            2025 Apr 22:101269. doi: 10.1016/j.cels.2025.101269."
COMMENT     Sequence Label: EF1a-mGL-bGH
FEATURES             Location/Qualifiers
     source          1..4245
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      496..1084
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     985..1004
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      1249..1335
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1427..1454
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     1462..1485
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     promoter        1522..2703
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1752..2694
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2651..2671
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     2705..2729
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2759..2768
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2765..2767,2768..2770,2771..3484)
                     /codon_start=1
                     /product="monomeric GFP derivative that displays 
                     exceptionally high brightness in cells (Campbell et al., 
                     2020)"
                     /label=mGreenLantern
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLGYGVACFARYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIVLKGIDFKEDGNILGHKLEYNFNSHKVYITADKQKNGIK
                     ANFKTRHNVEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLK
                     ERVTAAGITHDMDELYK"
     primer_bind     complement(2810..2831)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    3496..3720
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3831..3850)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             join(3842..4245,1..406)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPGDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3909..3928)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 taattgtcct tttaacagcg atcgcgtatt tcgtctagct caggcgcaat cacgaatgaa
       61 taacggtttg gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca
      121 agtctggaaa gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg
      181 tgatttctca cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt
      241 tggacgagtc ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg
      301 tgagttttct ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga
      361 tatgaataaa ttgcagtttc atttgatgct cgatgagttt ttctaatcag aattggttaa
      421 ttggttgtaa cattattcag attgggcccc gttccactga gcgtcagacc ccgtagaaaa
      481 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
      541 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
      601 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
      661 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
      721 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
      781 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
      841 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
      901 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
      961 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     1021 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     1081 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     1141 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg ctagcatgga
     1201 tctcggggac gtctaactac taagcgagag tagggaactg ccaggcatca aataaaacga
     1261 aaggctcagt cggaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc
     1321 ctgagtagga caaatccgcc gggagcggat ttgaacgttg tgaagcaacg gcccggaggg
     1381 tggcgggcag gacgcccgcc ataaactgcc aggcatcaaa ctaagcagaa ggccatcctg
     1441 acggatggcc tttttgcgtt tctacaaact cttcctgtta gttagttact taagctcggg
     1501 cccacctgca gtccttatac tgctccggtg cccgtcagtg ggcagagcgc acatcgccca
     1561 cagtccccga gaagttgggg ggaggggtcg gcaattgaac cggtgcctag agaaggtggc
     1621 gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc gagggtgggg
     1681 gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg
     1741 ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg
     1801 gcccttgcgt gccttgaatt acttccacgc ccctggctgc agtacgtgat tcttgatccc
     1861 gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg agccccttcg
     1921 cctcgtgctt gagttgaggc ctggcttggg cgctggggcc gccgcgtgcg aatctggtgg
     1981 caccttcgcg cctgtctcgc tgctttcgat aagtctctag ccatttaaaa tttttgatga
     2041 cctgctgcga cgcttttttt ctggcaagat agtcttgtaa atgcgggcca agatctgcac
     2101 actggtattt cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc ccagcgcaca
     2161 tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg gtagtctcaa
     2221 gctggccggc ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc gccctgggcg
     2281 gcaaggctgg cccggtcggc accagttgcg tgagcggaaa gatggccgct tcccggccct
     2341 gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg tgagtcaccc
     2401 acacaaagga aaagggcctt tccgtcctca gccgtcgctt catgtgactc cacggagtac
     2461 cgggcgccgt ccaggcacct cgattagttc tcgagctttt ggagtacgtc gtctttaggt
     2521 tggggggagg ggttttatgc gatggagttt ccccacactg agtgggtgga gactgaagtt
     2581 aggccagctt ggcacttgat gtaattctcc ttggaatttg ccctttttga gtttggatct
     2641 tggttcattc tcaagcctca gacagtggtt caaagttttt ttcttccatt tcaggtgtcg
     2701 tgagagctcg tttagtgaac cgtcagatcg cctggagacg ccatccacgc tgtaatgagc
     2761 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     2821 ggacggcgac gtaaacggcc acaagttcag cgtccgcggc gagggcgagg gcgatgccac
     2881 caacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     2941 caccctcgtg accaccttag gctacggcgt ggcctgcttc gcccgctacc ccgaccacat
     3001 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     3061 ctctttcaag gacgacggta cctacaagac ccgcgccgag gtgaagttcg agggcgacac
     3121 cctggtgaac cgcatcgtgc tgaagggcat cgacttcaag gaggacggca acatcctggg
     3181 gcacaagctg gagtacaact tcaacagcca caaggtctat atcacggccg acaagcagaa
     3241 gaacggcatc aaggctaact tcaagacccg ccacaacgtt gaggacggcg gcgtgcagct
     3301 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     3361 ccactacctg agccatcagt ccaaactgag caaagacccc aacgagaagc gcgatcacat
     3421 ggtcctgaag gagagggtga ccgccgccgg gattacacat gacatggacg agctgtacaa
     3481 gtaacaacgt tgcacctgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     3541 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     3601 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     3661 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     3721 cgctcgctga ctgcaggtgc ccgtgtctca aaatctctga tgttacattg cacaagataa
     3781 aaatatatca tcatgaacaa taaaactgtc tgcttacata aacagtaata caaggggtgt
     3841 tatgagccat attcaacggg aaacgtcgag gccgcgatta aattccaaca tggatgctga
     3901 tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg
     3961 cttgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag gtagcgttgc
     4021 caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta tgcctcttcc
     4081 gaccatcaag cattttatcc gtactcctgg tgatgcatgg ttactcacca ctgcgatccc
     4141 cggaaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga
     4201 tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttg
//
