LOCUS       sequence-191808-        5478 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5478)
  TITLE     C-terminal fluorescent fusion vectors for E. coli expression
REFERENCE   2  (bases 1 to 5478)
  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"
COMMENT     Sequence Label: pET GFP LIC cloning vector (u-msfGFP)
FEATURES             Location/Qualifiers
     source          1..5478
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     535..556
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(722..740)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      736..783
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(1149..1154,1155..1171,1172..1177))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1204..1222)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1290..1394
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1395..1463,1464..2255)
                     /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(1613..1632)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2426..3014
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2915..2934
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3168..3185
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3200..3339
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3265..3284)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3425..3447
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3441..3632)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5090..5109
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5193..5212
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5193..5211
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             5300..5322
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             join(5363..5365,5366..5478,1..601)
                     /codon_start=1
                     /product="enhanced GFP (EGFP) derivative that exhibits 
                     robust folding and enhanced photostability (Zhong et al., 
                     2019)"
                     /label=SiriusGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5405..5426)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
       61 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
      121 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctc
      181 cttcaaggac gacggcacct acaagacccg cgccgaggtg aagttcgagg gcgacaccct
      241 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
      301 caagctggag tacaacttca acagccacaa cgtctatatc acggccgaca agcagaagaa
      361 cggcatcaag gcgaacttca agatccgcca caacgtcgag gacggcagcg tgcagctcgc
      421 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
      481 ctacctgagc acccagtcca agctgagcaa agaccccaac gagaagcgcg atcacatggt
      541 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta
      601 gtaaccaact ccataaggat ccgcgatcgc ggcgcgccac ctggtggccg gccggtacca
      661 cgcgtgcgcg ctgatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
      721 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
      781 ttgctgaaag gaggaactat atccggacat ccacaggacg ggtgtggtcg ccatgatcgc
      841 gtagtcgata gtggctccaa gtagcgaagc gagcaggact gggcggcggc caaagcggtc
      901 ggacagtgct ccgagaacgg gtgcgcatag aaattgcatc aacgcatata gcgctagcag
      961 cacgccatag tgactggcga tgctgtcgga atggacgaca tcccgcaaga ggcccggcag
     1021 taccggcata accaagccta tgcctacagc atccagggtg acggtgccga ggatgacgat
     1081 gagcgcattg ttagatttca tacacggtgc ctgactgcgt tagcaattta actgtgataa
     1141 actaccgcat taaagcttat cgatgataag ctgtcaaaca tgagaattct tgaagacgaa
     1201 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     1261 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     1321 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataacatt
     1381 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     1441 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     1501 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     1561 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     1621 gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt
     1681 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     1741 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     1801 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     1861 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     1921 gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac
     1981 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     2041 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     2101 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     2161 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     2221 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     2281 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     2341 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     2401 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     2461 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     2521 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     2581 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     2641 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     2701 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     2761 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     2821 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     2881 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     2941 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     3001 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     3061 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     3121 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     3181 gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     3241 cacaccgcaa tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta
     3301 tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc
     3361 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     3421 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag
     3481 ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc
     3541 gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc
     3601 atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg
     3661 ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat
     3721 gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg
     3781 cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt
     3841 gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc
     3901 gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt
     3961 gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat
     4021 tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc
     4081 acgatcatgc gcacccgtgg ccaggaccca acgctgcccg agatgcgccg cgtgcggctg
     4141 ctggagatgg cggacgcgat ggatatgttc tgccaagggt tggtttgcgc attcacagtt
     4201 ctccgcaaga attgattggc tccaattctt ggagtggtga atccgttagc gaggtgccgc
     4261 cggcttccat tcaggtcgag gtggcccggc tccatgcacc gcgacgcaac gcggggaggc
     4321 agacaaggta tagggcggcg cctacaatcc atgccaaccc gttccatgtg ctcgccgagg
     4381 cggcataaat cgccgtgacg atcagcggtc caatgatcga agttaggctg gtaagagccg
     4441 cgagcgatcc ttgaagctgt ccctgatggt cgtcatctac ctgcctggac agcatggcct
     4501 gcaacgcggg catcccgatg ccgccggaag cgagaagaat cataatgggg aaggccatcc
     4561 agcctcgcgt cgcgaacgcc agcaagacgt agcccagcgc gtcggccgcc atgccggcga
     4621 taatggcctg cttctcgccg aaacgtttgg tggcgggacc agtgacgaag gcttgagcga
     4681 gggcgtgcaa gattccgaat accgcaagcg acaggccgat catcgtcgcg ctccagcgaa
     4741 agcggtcctc gccgaaaatg acccagagcg ctgccggcac ctgtcctacg agttgcatga
     4801 taaagaagac agtcataagt gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc
     4861 tgactgggtt gaaggctctc aagggcatcg gtcgacgctc tcccttatgc gactcctgca
     4921 ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg
     4981 catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc atacccacgc
     5041 cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg
     5101 cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg atgcgtccgg
     5161 cgtagaggat cgagatctcg atcccgcgaa attaatacga ctcactatag ggagaccaca
     5221 acggtttccc tctagtgccg gctccggaga gctctttaat taagcggccg ccctgcagga
     5281 ctcgagttct agaaataatt ttgtttaact ttaagaagga gatatagata tcctctcatc
     5341 ctccaacggc agcagcggat ccgtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     5401 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgc gcggcgaggg
     5461 cgagggcgat gccaccaa
//
