LOCUS       sequence-156016-        6260 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  an updated version of cLac94 (pACSA_PcptOO_YFP_PMB2__LacIWF). The 
            homologous region upstream of the inducible system was fixed to add 
            a stop codon for A1837. No difference in expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6260)
  AUTHORS   Markley AL, Begemann MB, Clarke RE, Gordon GC, Pfleger BF
  TITLE     Synthetic biology toolbox for controlling gene expression in the 
            cyanobacterium Synechococcus sp. strain PCC 7002.
  JOURNAL   ACS Synth Biol. 2015 May 15;4(5):595-603. doi: 10.1021/sb500260k. 
            Epub 2014 Sep 25.
  PUBMED    25216157
REFERENCE   2  (bases 1 to 6260)
  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: "10.1021/sb500260k";
            journalName: "ACS Synth Biol"; date: "2015-05-15- 15"; volume: "4"; 
            issue: "5"; pages: "595-603"
COMMENT     Sequence Label: pALM174
FEATURES             Location/Qualifiers
     source          1..6260
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(64..83)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     274..295
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        510..614
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(615..683,684..1475)
                     /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(833..852)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1646..2234
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2135..2154
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2388..2405
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3117..3136
                     /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."
     protein_bind    3221..3237
                     /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)."
     CDS             3286..3315
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             3328..3339
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             join(3361..3363,3364..3366,3367..4080)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3406..3427)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3667..3686)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4014..4035
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4151..4169)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4165..4212
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        4310..4344
                     /label=BBa_J23110
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.33"
     CDS             4374..5456
                     /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
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDFSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4393..4412)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     primer_bind     complement(6054..6073)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6055..6073)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6080..6097)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6080..6096)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6089..6111)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      join(6237..6260,1..432)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
       61 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
      121 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
      181 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
      241 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
      301 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
      361 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
      421 cgcttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
      481 catcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
      541 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
      601 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
      661 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
      721 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
      781 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
      841 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
      901 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
      961 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     1021 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     1081 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     1141 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     1201 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     1261 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     1321 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     1381 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     1441 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     1501 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     1561 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     1621 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     1681 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     1741 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     1801 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     1861 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     1921 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     1981 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     2041 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     2101 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     2161 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     2221 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     2281 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     2341 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     2401 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     2461 attaatgcag ctggcacgac aggtttcccg actggaactg ccgcactcaa tgttcaacca
     2521 gatctagtga ttatttcggg gttagcagcc gatgcgggca acctggtgaa gcaactgcga
     2581 gaattaggtt acaacggcat tattgttggg gggaacggct taaatacttc taatattttc
     2641 cccgtctgcc aagcaaaatg tgatggggtg ttggtggccc aagcctacag tgccgagtta
     2701 gataatgaga ttaaccgcgc gtttcgggac gcctattttc aacaaaacca aaaagagccg
     2761 ccccaattta gtgcccaggc ttttacggcg atccaagttt ttgttgaagc cctcagcagc
     2821 ctcgatgaaa aaacgccctt agaaactctt gctctaccgg acttgcgacg acaactgcgg
     2881 gacgaaattt ttgcaggtac ctacgtcacg cctttgggtg aaatttcctt cacagaggaa
     2941 ggggaaattg tccagaagga attttttgtg gcccaaattg aaatggatga atcgggtcaa
     3001 caggggcgtt tcgccttcat tgaaacgaac taaagtcaaa cctaggcctg tgtttaaaag
     3061 ggggaatcag ttttgtcttt ggaatttact gactcccctc tggacatctc caaacgaatt
     3121 gtgagcgctc acaattcgga attcttaaca aaaaagcagg aataaaatta acaagatgta
     3181 acagacataa gtcccatcac cgttgtataa atgtgtggaa ttgtgagcgg ataacaattt
     3241 cacacaccaa ctcataaagt caagtaggag attaattcca tgggccatca tcatcatcat
     3301 catcatcatc atcacagcag cggccatatc gaaggtcgtc atatgcaggc agcagcctcg
     3361 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     3421 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     3481 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     3541 ctcgtgacca ccttcggcta cggcctgcag tgcttcgccc gctaccccga ccacatgaag
     3601 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     3661 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     3721 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     3781 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     3841 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     3901 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     3961 tacctgagct accagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     4021 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
     4081 cgagggcggt gctttggcag gatccggctg tctaacaaag cccgaaagga agctgagttg
     4141 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
     4201 acgggttttt tgtctagatc aacggcctca acctactact gggctgcttc ctaatgcagg
     4261 agtcgcataa gggagagcgt cgagatcccg gacaccatcg aatggcgcat ttacagctag
     4321 ctcagtccta ggtacaatgc tagccggagg agggtcaatt catggtggtg aatgtgaaac
     4381 cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg
     4441 tggtgaacca ggccagccac gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg
     4501 cggagctgaa ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc
     4561 tgattggcgt tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga
     4621 ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg
     4681 gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga
     4741 tcattaacta tccgctggat gaccaggatg ccattgctgt ggaagctgcc tgcactaatg
     4801 ttccggcgtt atttcttgat gtctctgacc agacacccat caacagtatt attttctccc
     4861 atgaagacgg tacgcgactg ggcgtggagc atctggtcgc attgggtcac cagcaaatcg
     4921 cgctgttagc gggcccatta agttctgtct cggcgcgtct gcgtctggct ggctggcata
     4981 aatatctcac tcgcaatcaa attcagccga tagcggaacg ggaaggcgac tttagtgcca
     5041 tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc actgcgatgc
     5101 tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc cattaccgag tccgggctgc
     5161 gcgttggtgc ggatatctcg gtagtgggat acgacgatac cgaagacagc tcatgttata
     5221 tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc
     5281 gcttgctgca actctctcag ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac
     5341 tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg
     5401 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     5461 aacgcaatta atgtaagtta gctcactcat taggcaccgg gatctcgacc gatgcccttg
     5521 agagccttca acccagtcag ctccttccgg ttcgcgctca agttgatcga atgtatgccc
     5581 cggatatgac ctagggagat tcttctaaag cgagcagggt ttgccaagtg gcgatcgcct
     5641 cagggaacca taaccaactt tgttcgaggc ggcgggggtt gagctgggaa cgatcgagat
     5701 ctagggcgat cgcccgtagg gaaagccgaa aaactccaag aatttcctgg tcaacgtcgg
     5761 tgggttgcct ttgggccaaa tcgtacatac taatcgccag gcccgcataa atctccgcct
     5821 ggagggctgg ggggaaatcc tgggtcgttg ccgccgctaa ggcttcatac caacgatcat
     5881 tggcggcttc gtaatctgca ttgaggtaat ggacaaaacc cagggccatc aaaatttcgg
     5941 ggtcgttggg ctgttgggct agggccatgt cccaattgtc ttgcaccgtg gcgatcgccg
     6001 ttgcttgggt tggcgatggt gggacggcag ccaaagttgt ttgccagcat gcgccctata
     6061 gtgagtcgta ttacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     6121 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     6181 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggacgc
     6241 gccctgtagc ggcgcattaa
//
