LOCUS       sequence-386179-        5600 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  p15A ori, lacI, Ptrc promoter, AmpR. Kringle YFP (Yellow 
            Fluorescence Protein) expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5600)
  AUTHORS   Lozano Terol G, Gallego-Jara J, Sola Martinez RA, Martinez Vivancos 
            A, Canovas Diaz M, de Diego Puente T
  TITLE     Impact of the Expression System on Recombinant Protein Production in
            Escherichia coli BL21.
  JOURNAL   Front Microbiol. 2021 Jun 21;12:682001. doi: 
            10.3389/fmicb.2021.682001. eCollection 2021.
  PUBMED    34234760
REFERENCE   2  (bases 1 to 5600)
  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: "Front 
            Microbiol."; date: "2021-06-21"; volume: "12:682001. doi"; pages: " 
            10.3389/fmicb.2021.682001. eCollection 2021"
COMMENT     Sequence Label: pSF-p15A-trc- YFP
FEATURES             Location/Qualifiers
     source          1..5600
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    18..89
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(94..140)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    158..206
                     /note="synthetic polyadenylation signal"
     polyA_signal    241..289
                     /note="synthetic polyadenylation signal"
     misc_feature    303..394
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     343..362
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        401..478
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             479..1561
                     /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
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(498..517)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1793..1798,1799..1815,1816..1822)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1830..1846
                     /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)."
     polyA_signal    2689..2810
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2700..2719)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2754..2773
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2925..2971
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      3001..3137
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3182..3253
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3348..3893)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(3428..3447)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(4457..4525,4526..5317)
                     /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(4675..4694)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5330..5466
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
ORIGIN
        1 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt
       61 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga
      121 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc
      181 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctgat
      241 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgaaagta
      301 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      361 cccagtgcaa atgcaggtgc cagaacattt ctctgtcgac gacaccatcg aatggtgcaa
      421 aacctttcgc ggtatggcat gatagcgccc ggaagagagt caattcaggg tggtgaatgt
      481 gaaaccagta acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc
      541 ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc
      601 gatggcggag ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc
      661 gttgctgatt ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc
      721 ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg
      781 aagcggcgtc gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg
      841 gctgatcatt aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac
      901 taatgttccg gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt
      961 ctcccatgaa gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca
     1021 aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg
     1081 gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag
     1141 tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc
     1201 gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg
     1261 gctgcgcgtt ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg
     1321 ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt
     1381 ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt
     1441 ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc
     1501 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     1561 agcgcaacgc aattaatgta agttagcgcg aattgatctg gtttgacagc ttatcatcga
     1621 ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg gtatggctgt
     1681 gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt tctggataat
     1741 gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc tgttgacaat
     1801 taatcatccg gctcgtataa tgtgtggaat tgtgagcgga taacaagaat tcaaaggagg
     1861 tacccaccat gacggcactg actgaaggcg caaaactgtt cgagaaagaa atcccatata
     1921 tcactgagct ggaaggtgac gttgaaggta tgaagtttat catcaagggt gaaggcaccg
     1981 gtgacgcgag cgtcggtaaa gtggatgctc agttcatttg taccacgggc gacgttccgg
     2041 ttccgtggag cacgctggtc accacgctga cgtatggtgc tcagtgcttt gccaagtatc
     2101 cgcgccacat tgcggatttc ttcaaaagct gcatgccgga aggttacgtc caagagcgca
     2161 ccatcacctt tgagggtgat ggcgtgttca agacccgtgc ggaagtcacc tttgaaaatg
     2221 gcagcgtgta caaccgtgta aaactgaacg gccagggttt caagaaggac ggccacgtgc
     2281 tgggcaaaaa tctggagttt aactttaccc ctcattgttt gtacatttgg ggtgaccaag
     2341 cgaatcatgg cctgaagagc gcgttcaaaa tcatgcatga gatcaccggc tccaaagagg
     2401 atttcattgt tgccgatcac acccaaatga ataccccgat tggtggtggt ccggtgcacg
     2461 tgccggagta ccaccacatt acgtatcatg ttaccctgtc taaagacgtc accgatcacc
     2521 gtgaccattt gaacattgtt gaggtgatca aggcagttga cctggagacg taccgctaga
     2581 agttgtctcc tcctgcactg actgactgat acaatcgatt tctggatccg caggcctctg
     2641 ctagcttgac tgactgagat acagcgtacc ttcagctcac agacatgata agatacattg
     2701 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
     2761 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca
     2821 attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt
     2881 aaaacctcta caaatgtggt attggcccat ctctatcggt atcgtagcat aaccccttgg
     2941 ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg ggccggattg ctatctaccg
     3001 gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc
     3061 cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac
     3121 cagaaattta tccttaaggt cgtcagctat cctgcaggcg atctctcgat ttcgatcaag
     3181 acattccttt aatggtcttt tctggacacc actaggggtc agaagtagtt catcaaactt
     3241 tcttccctcc ctaatctcat tggttacctt gggctatcga aacttaatta aggaagatgc
     3301 caggaagata cttaacaggg aagtgagagg gccgcggcaa agccgttttt ccataggctc
     3361 cgcccccctg acaagcatca cgaaatctga cgctcaaatc agtggtggcg aaacccgaca
     3421 ggactataaa gataccaggc gtttccccct ggcggctccc tcgtgcgctc tcctgttcct
     3481 gcctttcggt ttaccggtgt cattccgctg ttatggccgc gtttgtctca ttccacgcct
     3541 gacactcagt tccgggtagg cagttcgctc caagctggac tgtatgcacg aaccccccgt
     3601 tcagtccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggaaagaca
     3661 tgcaaaagca ccactggcag cagccactgg taattgattt agaggagtta gtcttgaagt
     3721 catgcgccgg ttaaggctaa actgaaagga caagttttgg tgactgcgct cctccaagcc
     3781 agttacctcg gttcaaagag ttggtagctc agagaacctt cgaaaaaccg ccctgcaagg
     3841 cggttttttc gttttcagag caagagatta cgcgcagacc aaaacgatct caagaagatc
     3901 atcttattaa ggggtctgac gctcagtgga acgaaaaatt tggtggtatt taaatttccg
     3961 aactctccaa ggccctcgtc ggaaaatctt caaacctttc gtccgatcca tcttgcaggc
     4021 tacctctcga acgaactatc gcaagtctct tggccggcct tgcgccttgg ctattgcttg
     4081 gcagcgccta tcgccaggta ttactccaat cccgaatatc cgagatcggg atcacccgag
     4141 agaagttcaa cctacatcct caatcccgat ctatccgaga tccgaggaat atcgaaatcg
     4201 gggcgcgcct ggtgtaccga gaacgatcct ctcagtgcga gtctcgacga tccatatcgt
     4261 tgcttggcag tcagccagtc ggaatccagc ttgggaccca ggaagtccaa tcgtcagata
     4321 ttgtactcaa gcctggtcac ggcagcgtac cgatctgttt aaacctagat attgatagtc
     4381 tgatcggtca acgtataatc gagtcctagc ttttgcaaac atctatcaag agacaggatc
     4441 agcaggaggc tttcgcatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     4501 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     4561 agatcagttg ggtgcgcgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     4621 tgagagtttt cgccccgaag aacgctttcc aatgatgagc acttttaaag ttctgctatg
     4681 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     4741 ttctcagaat gacttggttg agtattcacc agtcacagaa aagcatctta cggatggcat
     4801 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     4861 acttctgaca acgattggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     4921 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     4981 gcgtgacacc acgatgcctg tagcaatggc aacaaccttg cgtaaactat taactggcga
     5041 actacttact ctagcttccc ggcaacagtt gatagactgg atggaggcgg ataaagttgc
     5101 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     5161 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     5221 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     5281 cgctgagata ggtgcctcac tgattaagca ttggtaaccg attctaggtg cattggcgca
     5341 gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc cacatatgcc agattcagca
     5401 acggatacgg cttccccaac ttgcccactt ccatacgtgt cctccttacc agaaatttat
     5461 ccttaagatc ccgaatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga
     5521 gcttacgcga acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc
     5581 gtcagcttac ctttttggca
//
