LOCUS       sequence-7050-ex        4939 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 4939)
  AUTHORS   Colwill K, Wells CD, Elder K, Goudreault M, Hersi K, Kulkarni S, 
            Hardy WR, Pawson T, Morin GB
  TITLE     Modification of the Creator recombination system for proteomics 
            applications--improved expression by addition of splice sites.
  JOURNAL   BMC Biotechnol. 2006 Mar 6;6:13. doi: 10.1186/1472-6750-6-13.
  PUBMED    16519801
REFERENCE   2  (bases 1 to 4939)
  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: "BMC 
            Biotechnol."; date: "2006-03-6"; volume: "6:13. doi"; pages: " 
            10.1186/1472-6750-6-13"
COMMENT     Sequence Label: V1579 pLP ProHis HTb - Acceptor
FEATURES             Location/Qualifiers
     source          1..4939
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    230..246
                     /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)."
     primer_bind     235..257
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             279..296
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             318..338
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     protein_bind    complement(351..384)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(633..650)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      803..889
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      981..1008
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1028..1119
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1120..1188,1189..1980)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPTAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1338..1357)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2151..2739
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2640..2659
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2893..2910
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2987..3415)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3097..3118)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3309..3328
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(3453..3472)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3640..3662
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        3740..3817
                     /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             3818..4900
                     /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(3837..3856)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gtttgacagc ttatcatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc
       61 ggaagctgtg gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc
      121 gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc
      181 tgaaatgagc tgttgacaat taatcatccg gtccgtataa tctgtggaat tgtgagcgga
      241 taacaatttc acacaggaaa cagaccatgt cgtactacca tcaccatcac catcacgatt
      301 acgatatccc aacgaccgaa aacctgtatt ttcagggcgc catgggatcc ataacttcgt
      361 atagcataca ttatacgaag ttatagatcc aatattattg aagcatttat cagggttatt
      421 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
      481 gcacatttcc ccgaaaagtg ccacctgacg tgaattcaaa ggcctacgtc gacgagctca
      541 ctagtcgcgg ccgctttcga atctagagcc tgcagtctcg aggcatgcgg taccaagctt
      601 ggctgttttg gcggatgaga gaagattttc agcctgatac agattaaatc agaacgcaga
      661 agcggtctga taaaacagaa tttgcctggc ggcagtagcg cggtggtccc acctgacccc
      721 atgccgaact cagaagtgaa acgccgtagc gccgatggta gtgtggggtc tccccatgcg
      781 agagtaggga actgccaggc atcaaataaa acgaaaggct cagtcgaaag actgggcctt
      841 tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc cgccgggagc
      901 ggatttgaac gttgcgaagc aacggcccgg agggtggcgg gcaggacgcc cgccataaac
      961 tgccaggcat caaattaagc agaaggccat cctgacggat ggcctttttg cgtttctaca
     1021 aactcttttt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     1081 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     1141 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     1201 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     1261 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     1321 agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag
     1381 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     1441 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     1501 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     1561 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     1621 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctacagcaat ggcaacaacg
     1681 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     1741 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     1801 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     1861 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     1921 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     1981 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     2041 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     2101 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     2161 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     2221 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     2281 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     2341 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2401 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2461 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2521 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2581 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2641 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2701 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2761 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     2821 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     2881 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     2941 ctccttacgc atctgtgcgg tatttcacac cgcataattt tgttaaaatt cgcgttaaat
     3001 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     3061 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     3121 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     3181 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     3241 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     3301 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     3361 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat
     3421 tcgccattca ggctgctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3481 agccagtacc agtcacgtag cgatatcgga gtgtatacac tccgctatcg ctacgtgact
     3541 gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     3601 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     3661 ggttttcacc gtcatcaccg aaacgcgcga ggcagcagat caattcgcgc gcgaaggcga
     3721 agcggcatgc atttacgttg acaccatcga atggtgcaaa acctttcgcg gtatggcatg
     3781 atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa cgttatacga
     3841 tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag
     3901 ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat
     3961 tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac
     4021 ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga
     4081 tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa
     4141 agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct
     4201 ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct
     4261 tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg
     4321 actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc
     4381 attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa
     4441 tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca
     4501 aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca
     4561 gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggatat
     4621 ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac
     4681 catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc
     4741 tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac
     4801 caccctggca cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     4861 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     4921 gttagcgcga attgatctg
//
