LOCUS       sequence-384159-        5688 bp    DNA     circular SYN 15-APR-2025
DEFINITION  p15A ori, lacI PT7 promoter, AmpR. Kringle YFP (Yellow Fluorescence 
            Protein) expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5688)
  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 5688)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-T7- YFP
FEATURES             Location/Qualifiers
     source          1..5688
                     /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"
     CDS             complement(401..1483)
                     /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     1445..1464
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1484..1561)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1767..1786
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1870..1889
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1870..1888
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1889..1913
                     /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    2777..2898
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2788..2807)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2842..2861
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      3013..3059
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      3089..3225
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3270..3341
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3436..3981)
                     /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(3516..3535)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(4545..4613,4614..5405)
                     /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(4763..4782)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5418..5554
                     /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 tcactgcccg ctttccagtc
      421 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
      481 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
      541 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
      601 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
      661 cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc
      721 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
      781 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
      841 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
      901 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
      961 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
     1021 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
     1081 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
     1141 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
     1201 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
     1261 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
     1321 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
     1381 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
     1441 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     1501 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     1561 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     1621 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     1681 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     1741 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     1801 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc
     1861 ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctaa
     1921 gcttccgagc tctcgaattc aaaggaggta cccaccatga cggcactgac tgaaggcgca
     1981 aaactgttcg agaaagaaat cccatatatc actgagctgg aaggtgacgt tgaaggtatg
     2041 aagtttatca tcaagggtga aggcaccggt gacgcgagcg tcggtaaagt ggatgctcag
     2101 ttcatttgta ccacgggcga cgttccggtt ccgtggagca cgctggtcac cacgctgacg
     2161 tatggtgctc agtgctttgc caagtatccg cgccacattg cggatttctt caaaagctgc
     2221 atgccggaag gttacgtcca agagcgcacc atcacctttg agggtgatgg cgtgttcaag
     2281 acccgtgcgg aagtcacctt tgaaaatggc agcgtgtaca accgtgtaaa actgaacggc
     2341 cagggtttca agaaggacgg ccacgtgctg ggcaaaaatc tggagtttaa ctttacccct
     2401 cattgtttgt acatttgggg tgaccaagcg aatcatggcc tgaagagcgc gttcaaaatc
     2461 atgcatgaga tcaccggctc caaagaggat ttcattgttg ccgatcacac ccaaatgaat
     2521 accccgattg gtggtggtcc ggtgcacgtg ccggagtacc accacattac gtatcatgtt
     2581 accctgtcta aagacgtcac cgatcaccgt gaccatttga acattgttga ggtgatcaag
     2641 gcagttgacc tggagacgta ccgctagaag ttgtctcctc ctgcactgac tgactgatac
     2701 aatcgatttc tggatccgca ggcctctgct agcttgactg actgagatac agcgtacctt
     2761 cagctcacag acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag
     2821 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     2881 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     2941 gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat tggcccatct
     3001 ctatcggtat cgtagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgt
     3061 gcccctcggg ccggattgct atctaccggc attggcgcag aaaaaaatgc ctgatgcgac
     3121 gctgcgcgtc ttatactccc acatatgcca gattcagcaa cggatacggc ttccccaact
     3181 tgcccacttc catacgtgtc ctccttacca gaaatttatc cttaaggtcg tcagctatcc
     3241 tgcaggcgat ctctcgattt cgatcaagac attcctttaa tggtcttttc tggacaccac
     3301 taggggtcag aagtagttca tcaaactttc ttccctccct aatctcattg gttaccttgg
     3361 gctatcgaaa cttaattaag gaagatgcca ggaagatact taacagggaa gtgagagggc
     3421 cgcggcaaag ccgtttttcc ataggctccg cccccctgac aagcatcacg aaatctgacg
     3481 ctcaaatcag tggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     3541 cggctccctc gtgcgctctc ctgttcctgc ctttcggttt accggtgtca ttccgctgtt
     3601 atggccgcgt ttgtctcatt ccacgcctga cactcagttc cgggtaggca gttcgctcca
     3661 agctggactg tatgcacgaa ccccccgttc agtccgaccg ctgcgcctta tccggtaact
     3721 atcgtcttga gtccaacccg gaaagacatg caaaagcacc actggcagca gccactggta
     3781 attgatttag aggagttagt cttgaagtca tgcgccggtt aaggctaaac tgaaaggaca
     3841 agttttggtg actgcgctcc tccaagccag ttacctcggt tcaaagagtt ggtagctcag
     3901 agaaccttcg aaaaaccgcc ctgcaaggcg gttttttcgt tttcagagca agagattacg
     3961 cgcagaccaa aacgatctca agaagatcat cttattaagg ggtctgacgc tcagtggaac
     4021 gaaaaatttg gtggtattta aatttccgaa ctctccaagg ccctcgtcgg aaaatcttca
     4081 aacctttcgt ccgatccatc ttgcaggcta cctctcgaac gaactatcgc aagtctcttg
     4141 gccggccttg cgccttggct attgcttggc agcgcctatc gccaggtatt actccaatcc
     4201 cgaatatccg agatcgggat cacccgagag aagttcaacc tacatcctca atcccgatct
     4261 atccgagatc cgaggaatat cgaaatcggg gcgcgcctgg tgtaccgaga acgatcctct
     4321 cagtgcgagt ctcgacgatc catatcgttg cttggcagtc agccagtcgg aatccagctt
     4381 gggacccagg aagtccaatc gtcagatatt gtactcaagc ctggtcacgg cagcgtaccg
     4441 atctgtttaa acctagatat tgatagtctg atcggtcaac gtataatcga gtcctagctt
     4501 ttgcaaacat ctatcaagag acaggatcag caggaggctt tcgcatgagt attcaacatt
     4561 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     4621 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcgcgagtg ggttacatcg
     4681 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgctttccaa
     4741 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     4801 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tattcaccag
     4861 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     4921 ccatgagtga taacactgcg gccaacttac ttctgacaac gattggagga ccgaaggagc
     4981 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     5041 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     5101 caaccttgcg taaactatta actggcgaac tacttactct agcttcccgg caacagttga
     5161 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     5221 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     5281 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     5341 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     5401 ggtaaccgat tctaggtgca ttggcgcaga aaaaaatgcc tgatgcgacg ctgcgcgtct
     5461 tatactccca catatgccag attcagcaac ggatacggct tccccaactt gcccacttcc
     5521 atacgtgtcc tccttaccag aaatttatcc ttaagatccc gaatcgttta aactcgactc
     5581 tggctctatc gaatctccgt cgtttcgagc ttacgcgaac agccgtggcg ctcatttgct
     5641 cgtcgggcat cgaatctcgt cagctatcgt cagcttacct ttttggca
//
