LOCUS       sequence-176159-        7000 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses  sfGFP fused to Cre in E.Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7000)
  TITLE     pET-sfGFP-GGS9-Cre-6xHis
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7000)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pET-sfGFP-GGS9-Cre-6xHis
FEATURES             Location/Qualifiers
     source          1..7000
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(43..631)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(123..142)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        712..804
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(805..873,874..1665)
                     /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(1023..1042)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(1780..2235)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1917..1938)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2129..2148
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(2272..2319)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     2315..2333
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(2371..2415)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(2434..2451)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(2461..3486)
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="SNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKMLL
                     SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP
                     RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL
                     GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER
                     WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR
                     YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED
                     GD"
     RBS             4300..4322
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4337..4361
                     /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     complement(4361..4380)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4362..4380)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4468..4487)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        4693..4770
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4771..5853
                     /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(4790..4809)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             6425..6616
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(6610..6632)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    6718..6857
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     6773..6792
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6872..6889)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
       61 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
      121 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
      181 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
      241 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
      301 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
      361 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
      421 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
      481 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
      541 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
      601 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
      661 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgacct atttgtttat
      721 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
      781 aatcatgatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
      841 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
      901 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
      961 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     1021 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     1081 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     1141 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     1201 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     1261 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     1321 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     1381 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     1441 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     1501 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     1561 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     1621 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     1681 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     1741 aagatccttt ttgataatct catgccatcg tgccgccaga aattgtaaac gttaatattt
     1801 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     1861 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     1921 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     1981 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     2041 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     2101 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     2161 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     2221 cgctacaggg cgcgtcccat tcgccaatcc ggatatagtt cctcctttca gcaaaaaacc
     2281 cctcaagacc cgtttagagg ccccaagggg ttatgctagt tattgctcag cggtggcagc
     2341 agcctaggtt aattaagctg cgctagtaga cgagtccatg tgctggcgtt caaatttcgc
     2401 agcagcggtt tctttaccag actcgagtca ttagtgatgg tggtggtgat gggatccgcc
     2461 gtcgccatct tccagcaggc gcaccattgc ccctgtttca ctatccaggt tacggatata
     2521 gttcatgaca atatttacat tggtccagcc accagcttgc atgatctccg gtattgaaac
     2581 tccagcgcgg gccatatctc gcgcggctcc gacacgggca ctgtgtccag accaggccag
     2641 gtatctctga ccagagtcat ccttagcgcc gtaaatcaat cgatgagttg cttcaaaaat
     2701 cccttccagg gcgcgagttg atagctggct ggtggcagat ggcgcggcaa caccattttt
     2761 tctgacccgg caaaacaggt agttattcgg atcatcagct acaccagaga cggaaatcca
     2821 tcgctcgacc agtttagtta cccccaggct aagtgccttc tctacacctg cggtgctaac
     2881 cagcgttttc gttctgccaa tatggattaa cattctccca ccgtcagtac gtgagatatc
     2941 tttaaccctg atcctggcaa tttcggctat acgtaacagg gtgttataag caatccccag
     3001 aaatgccaga ttacgtatat cctggcagcg atcgctattt tccatgagtg aacgaacctg
     3061 gtcgaaatca gtgcgttcga acgctagagc ctgttttgca cgttcaccgg catcaacgtt
     3121 ttcttttcgg atacgccgca taaccagtga aacagcattg ctgtcacttg gtcgtggcag
     3181 cccggaccga cgatgaagca tgtttagctg gcccaaatgt tgctggatag tttttactgc
     3241 cagaccgcgc gcctgaagat atagaagata atcgcgaaca tcttcaggtt ctgcgggaaa
     3301 ccatttccgg ttattcaact tgcaccatgc cgcccacgac cggcaaacgg acagaagcat
     3361 tttccaggta tgctcagaaa acgcctggcg atccctgaac atgtccatca ggttcttgcg
     3421 aacctcatca ctcgttgcat cgaccggcaa tgcaggcaaa ttttggtgta cggtcagtaa
     3481 attgctcgcg gtaccgccgc tgccaccgct accaccgcta ccaccgctac cgccgctacc
     3541 accagaacca ccagaaccac cgctaccacc ggtcttgtag agttcgtcca ttccgtgagt
     3601 aatgcctgca gcggtcacga attctaacag aaccatgtga tcgcgttttt cgttcggatc
     3661 tttgctcagt gctgactggg tgctcaggta gtggttgtca ggtaacagta caggcccgtc
     3721 accaatcggc gtgttttgtt gataatgatc cgctaattga acgctgccgt cttccacgtt
     3781 atggcgaatt ttgaaatttg ccttgatacc gtttttctgt ttatcggccg taatatacac
     3841 gttatgagaa ttaaagttat attccagctt atggcctaaa atgttgccgt cttctttgaa
     3901 gtcgatacct ttcagttcga tacggtttac taatgtgtcg ccttcaaatt tcacttccgc
     3961 acgcgtttta taggtgccat cgtccttgaa gctgattgta cgctcttgaa catatccttc
     4021 aggcatggct gatttgaaaa aatcgtgttg tttcatatga tctgggtaac gactgaagca
     4081 ctgaacgcca taggtcaggg tggtcactaa ggtgggccac ggaacgggta atttgcctgt
     4141 tgtgcaaata aacttcaggg tcaatttacc attggtagcg tcaccttcgc cttcgccgcg
     4201 cacgctaaat ttgtgaccgt tcacgtcgcc gtctaattcc actaagatcg gtactacacc
     4261 cgtaaacagc tcttcacctt tgctagcacc catggtatat ctccttctta aagttaaaca
     4321 aaattatttc tagaggggaa ttgttatccg ctcacaattc ccctatagtg agtcgtatta
     4381 atttcgcggg atcgagatcg atctcgatcc tctacgccgg acgcatcgtg gccggcatca
     4441 ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat ggggaagatc
     4501 gggctcgcca cttcgggctc atgagcgctt gtttcggcgt gggtatggtg gcaggccccg
     4561 tggccggggg actgttgggc gccatctcct tgcatgcacc attccttgcg gcggcggtgc
     4621 tcaacggcct caacctacta ctgggctgct tcctaatgca ggagtcgcat aagggagagc
     4681 gtcgagatcc cggacaccat cgaatggcgc aaaacctttc gcggtatggc atgatagcgc
     4741 ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca
     4801 gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt
     4861 tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac
     4921 cgcgtggcac aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt
     4981 ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg
     5041 ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg
     5101 gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac
     5161 caggatgcca ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc
     5221 tctgaccaga cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc
     5281 gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt
     5341 tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt
     5401 cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg
     5461 caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg
     5521 ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga catctcggta
     5581 gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa
     5641 caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc
     5701 caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg
     5761 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
     5821 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct
     5881 cactcattag gcaccgggat ctcgaccgat gcccttgaga gccttcaacc cagtcagctc
     5941 cttccggtgg gcgcggggca tgactagcat gatcgtgctc ctgtcgttga ggacccggct
     6001 aggctggcgg ggttgcctta ctggttagca gaatgaatca ccgatacgcg agcgaacgtg
     6061 aagcgactgc tgctgcaaaa cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt
     6121 ccgtgtttcg taaagtctgg aaacgcggaa gtcagcgccc tgcaccatta tgttccggat
     6181 ctgcatcgca ggatgctgct ggctaccctg tggaacacct acatctgtat taacgaagcg
     6241 ctggcattga ccctgagtga tttttctctg gtcccgccgc atccataccg ccagttgttt
     6301 accctcacaa cgttccagta accgggcatg ttcatcatca gtaacccgta tcgtgagcat
     6361 cctctctcgt ttcatcggta tcattacccc catgaacaga aatccccctt acacggaggc
     6421 atcagtgacc aaacaggaaa aaaccgccct taacatggcc cgctttatca gaagccagac
     6481 attaacgctt ctggagaaac tcaacgagct ggacgcggat gaacaggcag acatctgtga
     6541 atcgcttcac gaccacgctg atgagcttta ccgcagctgc ctcgcgcgtt tcggtgatga
     6601 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     6661 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc
     6721 agccatgacc cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca
     6781 gagcagattg tactgagagt gcaccattgc ggtgtgaaat accgcacaga tgcgtaagga
     6841 gaaaataccg catcaggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     6901 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     6961 caggggataa cgcaggaaag aacatgtgag caaaaggcca
//
