LOCUS       sequence-90574-e        4696 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP from jellyfish Aquorea sp. under T5 promoter with lac operators;
            ampicillin resistant..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4696)
  AUTHORS   Siddall A, Williams AA, Sanders J, Denton JA, Madden D, Schollar J, 
            Bryk J
  TITLE     UNIGEMS: plasmids and parts to facilitate teaching on assembly, gene
            expression control and logic in E. coli
  JOURNAL   bioRxiv 2021.06.20.449138
REFERENCE   2  (bases 1 to 4696)
  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: "bioRxiv 
            2021.06.20.449138"
COMMENT     Sequence Label: p007ampGFP
FEATURES             Location/Qualifiers
     source          1..4696
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      502..1090
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     991..1010
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1244..1261
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1359..1376
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1360..1376
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1832..1848
                     /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        join(1873..1887,1888..1893,1894..1910,1911..1917)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    1893..1909
                     /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)."
     protein_bind    1923..1942
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     RBS             1983..1991
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             1997..2713
                     /codon_start=1
                     /product="Aequorea victoria green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTRVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKA
                     NFKVRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     2650..2670
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(2909..2928)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(2934..2953)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        2995..3086
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3087..3155,3156..3947)
                     /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     complement(3305..3324)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             join(4030..4696,1..416)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAVVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4049..4068)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc actgcgatgc
       61 tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc cattaccgag tccgggctgc
      121 gcgttggtgc ggatatctcg gtagtgggat acgacgatac cgaagatagc tcatgttata
      181 tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc
      241 gcttgctgca actctctcag ggccaggcgg tgaagggcaa tcagctgttg ccagtctcac
      301 tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg
      361 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgactca
      421 tgaccaaaat cccttaacgt gagttacgcg cgcgtcgttc cactgagcgt cagaccccgt
      481 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
      541 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
      601 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
      661 gccgtagtta gcccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
      721 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
      781 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
      841 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
      901 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
      961 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     1021 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     1081 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     1141 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     1201 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     1261 ggaagcggaa ggcgagagta gggaactgcc aggcatcaaa ctaagcagaa ggcccctgac
     1321 ggatggcctt tttgcgtttc tacaaactct ttctgtgttg taaaacgacg gccagtctta
     1381 agctcgggcc ccctgggcgg ttctgataac gagtaatcgt taatccgcaa ataacgtaaa
     1441 aacccgcttc ggcgggtttt tttatggggg gagtttaggg aaagagcatt tgtcagaata
     1501 tttaagggcg cctgtcactt tgcttgatat atgagaatta tttaacctta taaatgagaa
     1561 aaaagcaacg cactttaaat aagatacgtt gctttttcga ttgatgaaca cctataatta
     1621 aactattcat ctattattta tgattttttg tatatacaat atttctagtt tgttaaagag
     1681 aattaagaaa ataaatctcg aaaataataa agggaaaatc agtttttgat atcaaaatta
     1741 tacatgtcaa cgataataca aaatataata caaactataa gatgttatca gtatttatta
     1801 tcatttagaa taaattttgt gtcgccctta attgtgagcg gataacaatt acgagcttca
     1861 tgcacagtga aatcatgaaa aatttatttg ctttgtgagc ggataacaat tataatatgt
     1921 ggaattgtga gcgctcacaa ttccacaacg gtttccctct agaaataatt ttgtttaact
     1981 tttaaggagg taaaaaatga gtaaaggaga agaacttttc actagagttg tcccaattct
     2041 tgttgaatta gacggtgatg ttaatgggca caaattttct gtcagtggag agggagaagg
     2101 tgatgcaaca tacggaaaac ttacccttaa atttatttgc actactggaa aactaccagt
     2161 tccgtggcca acacttgtca ctactctctc ttatggtgtt caatgctttt cgagataccc
     2221 agatcacatg aaacggcatg actttttcaa gagtgccatg cccgagggtt atgtacagga
     2281 aagaactata tttttcaaag atgacgggaa ctacaagaca cgtgctgaag tcaagtttga
     2341 aggtgatacc cttgttaata gaatcgagtt aaaaggtgtt gattttaaag aagatggaaa
     2401 cattcttgga cacaaattgg aatacaacta taactcacac aatgtataca tcatggcaga
     2461 caaacaaaag aatggaatca aagctaactt caaagttaga cacaacattg aagatggaag
     2521 cgttcaactg gcagaccatt atcaacaaaa tactccaatt ggcgatggcc ctgtcctttt
     2581 accagacaac cattacctgt ccacacaatc tgccctttcg aaagatccca acgaaaagag
     2641 agaccacatg gtccttcttg agtttgtaac agctgctggg attacacatg gcatggatga
     2701 actatacaaa taaccccaag ggcgacaccc cctaattagc ccgggcgaaa ggcccagtct
     2761 ttcgactgag cctttcgttt tatttgatgc ctggcagttc cctactctcg catggggagt
     2821 ccccacacta ccatcggcgc tacggcgttt cacttctgag ttcggcatgg ggtcaggtgg
     2881 gaccaccgcg ctactgccgc caggcaaaca aggggtgtta tgagccatat tcaggtataa
     2941 atgggctcgc gataatgttc agaattggtt aattggttgt aacactgacc cctatttgtt
     3001 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3061 ttcaataata ttgaaaaagg aagaatatga gtattcaaca tttccgtgtc gcccttattc
     3121 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3181 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3241 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3301 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     3361 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3421 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3481 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3541 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3601 caaacgacga gcgtgacacc acgatgcctg tagcgatggc aacaacgttg cgcaaactat
     3661 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     3721 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     3781 aatccggagc cggtgagcgt ggttctcgcg gtatcatcgc agcgctgggg ccagatggta
     3841 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     3901 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaagcg gcgcgccatc
     3961 gaatggcgca aaacctttcg cggtatggca tgatagcgcc cggaagagag tcaattcagg
     4021 gtggtgaata tgaaaccagt aacgttatac gatgtcgcag agtatgccgg tgtctcttat
     4081 cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac gcgggaaaaa
     4141 gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca acaactggcg
     4201 ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca cgcgccgtcg
     4261 caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt ggtggtgtcg
     4321 atggtagaac gaagcggcgt cgaagcctgt aaagcggtgg tgcacaatct tctcgcgcaa
     4381 cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat tgctgtggaa
     4441 gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac acccatcaac
     4501 agtattattt tctcccatga ggacggtacg cgactgggcg tggagcatct ggtcgcattg
     4561 ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc gcgtctgcgt
     4621 ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa
     4681 ggcgactgga gtgcca
//
