LOCUS       sequence-156568-        5374 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GFP with 3 UAG codons at the amino acid position 3, 151, 
            and 153.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5374)
  AUTHORS   Schwessinger B, Li X, Ellinghaus TL, Chan LJ, Wei T, Joe A, Thomas 
            N, Pruitt R, Adams PD, Chern MS, Petzold CJ, Liu CC, Ronald PC
  TITLE     A second-generation expression system for tyrosine-sulfated proteins
            and its application in crop protection.
  JOURNAL   Integr Biol (Camb). 2016 Apr 18;8(4):542-5. doi: 10.1039/c5ib00232j.
            Epub 2015 Nov 27.
  PUBMED    26611838
REFERENCE   2  (bases 1 to 5374)
  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"; doi: 
            "10.1039/c5ib00232j"; journalName: "Integr Biol (Camb)"; date: 
            "2016-04-18- 18"; volume: "8"; issue: "4"; pages: "542-5"
COMMENT     Sequence Label: pGLO-GFP-3UAG
FEATURES             Location/Qualifiers
     source          1..5374
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(96..974)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        1001..1285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     1208..1227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     RBS             1312..1334
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1351..2064
                     /codon_start=1
                     /product="Cycle 3 GFP (Crameri et al., 1996)"
                     /label=Cycle 3 GFP
                     /note="mammalian codon-optimized"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSH*V*ITADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(1378..1406)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2001..2021
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     misc_feature    2066..2122
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(2155..2172)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2324..2410
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2502..2529
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2547..2638
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2639..2707,2708..3499)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2857..2876)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3541..3996
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3628..3647)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3838..3859
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     rep_origin      4107..4695
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4596..4615
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4849..4866
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4881..5021
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4947..4966)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5107..5129
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5268..5287
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 atcgatgcat aatgtgcctg tcaaatggac gaagcaggga ttctgcaaac cctatgctac
       61 tccgtcaagc cgtcaattgt ctgattcgtt accaattatg acaacttgac ggctacatca
      121 ttcacttttt cttcacaacc ggcacggaac tcgctcgggc tggccccggt gcatttttta
      181 aatacccgcg agaaatagag ttgatcgtca aaaccaacat tgcgaccgac ggtggcgata
      241 ggcatccggg tggtgctcaa aagcagcttc gcctggctga tacgttggtc ctcgcgccag
      301 cttaagacgc taatccctaa ctgctggcgg aaaagatgtg acagacgcga cggcgacaag
      361 caaacatgct gtgcgacgct ggcgatatca aaattgctgt ctgccaggtg atcgctgatg
      421 tactgacaag cctcgcgtac ccgattatcc atcggtggat ggagcgactc gttaatcgct
      481 tccatgcgcc gcagtaacaa ttgctcaagc agatttatcg ccagcagctc cgaatagcgc
      541 ccttcccctt gcccggcgtt aatgatttgc ccaaacaggt cgctgaaatg cggctggtgc
      601 gcttcatccg ggcgaaagaa ccccgtattg gcaaatattg acggccagtt aagccattca
      661 tgccagtagg cgcgcggacg aaagtaaacc cactggtgat accattcgcg agcctccgga
      721 tgacgaccgt agtgatgaat ctctcctggc gggaacagca aaatatcacc cggtcggcaa
      781 acaaattctc gtccctgatt tttcaccacc ccctgaccgc gaatggtgag attgagaata
      841 taacctttca ttcccagcgg tcggtcgata aaaaaatcga gataaccgtt ggcctcaatc
      901 ggcgttaaac ccgccaccag atgggcatta aacgagtatc ccggcagcag gggatcattt
      961 tgcgcttcag ccatactttt catactcccg ccattcagag aagaaaccaa ttgtccatat
     1021 tgcatcagac attgccgtca ctgcgtcttt tactggctct tctcgctaac caaaccggta
     1081 accccgctta ttaaaagcat tctgtaacaa agcgggacca aagccatgac aaaaacgcgt
     1141 aacaaaagtg tctataatca cggcagaaaa gtccacattg attatttgca cggcgtcaca
     1201 ctttgctatg ccatagcatt tttatccata agattagcgg atcctacctg acgcttttta
     1261 tcgcaactct ctactgtttc tccatacccg tttttttggg ctagaaataa ttttgtttaa
     1321 ctttaagaag gagatataca tatggcttag agcaaaggag aagaactttt cactggagtt
     1381 gtcccaattc ttgttgaatt agatggtgat gttaatgggc acaaattttc tgtcagtgga
     1441 gagggtgaag gtgatgctac atacggaaag cttaccctta aatttatttg cactactgga
     1501 aaactacctg ttccatggcc aacacttgtc actactttct cttatggtgt tcaatgcttt
     1561 tcccgttatc cggatcatat gaaacggcat gactttttca agagtgccat gcccgaaggt
     1621 tatgtacagg aacgcactat atctttcaaa gatgacggga actacaagac gcgtgctgaa
     1681 gtcaagtttg aaggtgatac ccttgttaat cgtatcgagt taaaaggtat tgattttaaa
     1741 gaagatggaa acattctcgg acacaaactc gagtacaact ataactcaca ctaggtatag
     1801 atcacggcag acaaacaaaa gaatggaatc aaagctaact tcaaaattcg ccacaacatt
     1861 gaagatggat ccgttcaact agcagaccat tatcaacaaa atactccaat tggcgatggc
     1921 cctgtccttt taccagacaa ccattacctg tcgacacaat ctgccctttc gaaagatccc
     1981 aacgaaaagc gtgaccacat ggtccttctt gagtttgtaa ctgctgctgg gattacacat
     2041 ggcatggatg agctctacaa ataatgaatt cgagctcggt acccggggat cctctagagt
     2101 cgacctgcag gcatgcaagc ttggctgttt tggcggatga gagaagattt tcagcctgat
     2161 acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg gcggcagtag
     2221 cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta gcgccgatgg
     2281 tagtgtgggg tcccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc
     2341 tcagtgcaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag
     2401 taggacaaat ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg
     2461 ggcaggacgc ccgccataaa ctgccaggca tcaaattaag cagaaggcca tcctgacgga
     2521 tggccttttt gcgtttctac aaactctttg tttatttttc taaatacatt caaatatgta
     2581 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     2641 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     2701 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     2761 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     2821 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     2881 tgttgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     2941 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     3001 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     3061 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     3121 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     3181 tgcagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     3241 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     3301 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     3361 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     3421 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     3481 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     3541 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     3601 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     3661 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     3721 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     3781 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     3841 gactcttgtt ccaaactgga acaacactca accctatctc gggctattct tttgatttat
     3901 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     3961 acgcgaattt taacaaaata ttaacgttta caatttaaaa ggatctaggt gaagatcctt
     4021 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     4081 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     4141 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     4201 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     4261 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     4321 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     4381 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     4441 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcat
     4501 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     4561 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     4621 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     4681 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     4741 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     4801 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     4861 agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     4921 tcacaccgca tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     4981 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     5041 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     5101 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     5161 agcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg
     5221 aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
     5281 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
     5341 tagaggatct aattctcatg tttgacagct tatc
//
