LOCUS       sequence-343594-        4363 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template for making mRNA for overexpressing eGFP in frog oocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4363)
  AUTHORS   Xiang K, Bartel DP
  TITLE     The molecular basis of coupling between poly(A)-tail length and 
            translational efficiency.
  JOURNAL   Elife. 2021 Jul 2;10. pii: 66493. doi: 10.7554/eLife.66493.
  PUBMED    34213414
REFERENCE   2  (bases 1 to 4363)
  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: "Elife."; 
            date: "2021-07-2"; volume: "10. pii: 66493. doi"; pages: " 
            10.7554/eLife.66493"
COMMENT     Sequence Label: C079
FEATURES             Location/Qualifiers
     source          1..4363
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     5'UTR           16..58
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     CDS             join(70..72,73..75,76..789)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(70..72,73..75,76..786)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(115..136)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(376..395)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     723..744
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     3'UTR           789..914
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
     promoter        complement(1234..1252)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1235..1252)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1270..1286)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1270..1286)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1283..1305)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1294..1310
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1318..1324,1325..1342,1343..1348))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1363..1384
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1501..1518)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1672..2260)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1752..1771)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2431..3222,3223..3291))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3054..3073
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3292..3396)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3464..3482
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3520..3542)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3642..3661
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(3728..4183)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3865..3886)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4077..4096
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4309..4331
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4323..4340
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4324..4340
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(4347..4363,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(4347..4363,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaataca agcttcttgt tctttttgca gaagctcaga ataaacgctc aactttggca
       61 gatctgaaca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc
      121 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
      181 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
      241 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac
      301 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
      361 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
      421 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
      481 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
      541 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
      601 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
      661 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat
      721 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
      781 tacaagtaaa ccagcctcaa gaacacccga atggagtctc taagctacat aataccaact
      841 tacactttac aaaatgttgt cccccaaaat gtagccattc gtatctgctc ctaataaaaa
      901 gaaagtttct tcacattctg ggcgtagatc cagacatgaa aaaaaaaaaa aaaaaaaaaa
      961 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     1021 aaaaaaaaaa aaaaaaaaca gtagggttgt aaaggttttt cttttcctga gaaaacaacc
     1081 ttttgttttc tcaggttttg ctttttggcc tttccctagc tttaaaaaaa aaagcaaaag
     1141 ctagccatgg tcccagcctc ctcgctggcg gctagtgggc aacatgcttc ggcatggcga
     1201 atgggacgcg gccgcatgca taagcttgag tattctatag tgtcacctaa atagcttggc
     1261 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     1321 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     1381 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     1441 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     1501 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     1561 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     1621 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1681 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1741 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1801 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1861 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1921 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1981 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     2041 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     2101 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     2161 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     2221 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     2281 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     2341 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     2401 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     2461 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     2521 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     2581 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     2641 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     2701 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     2761 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     2821 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     2881 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     2941 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     3001 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     3061 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     3121 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     3181 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     3241 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     3301 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     3361 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     3421 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     3481 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     3541 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     3601 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
     3661 actgagagtg caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     3721 catcaggaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     3781 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     3841 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     3901 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     3961 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     4021 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     4081 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     4141 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtccattcg ccattcaggc
     4201 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     4261 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
     4321 gttgtaaaac gacggccagt gaattgtaat acgactcact ata
//
