LOCUS       sequence-133158-        7080 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  +36-GFP-Cre fusion expression in E.Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7080)
  AUTHORS   Li M, Tao Y, Shu Y, LaRochelle JR, Steinauer A, Thompson D, 
            Schepartz A, Chen ZY, Liu DR
  TITLE     Discovery and Characterization of a Peptide That Enhances Endosomal 
            Escape of Delivered Proteins in Vitro and in Vivo.
  JOURNAL   J Am Chem Soc. 2015 Nov 11;137(44):14084-93. doi: 
            10.1021/jacs.5b05694. Epub 2015 Oct 30.
  PUBMED    26465072
REFERENCE   2  (bases 1 to 7080)
  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/jacs.5b05694"; journalName: "J Am Chem Soc"; date: 
            "2015-11-11- 11"; volume: "137"; issue: "44"; pages: "14084-93"
COMMENT     Sequence Label: (GGS)9_(+36GFP)-Cre.ape
FEATURES             Location/Qualifiers
     source          1..7080
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    2..26
                     /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)."
     RBS             41..63
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             955..1980
                     /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"
     CDS             1990..2007
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2026..2070
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2108..2126)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2122..2169
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2206..2661
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2293..2312)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2503..2524
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2776..3567,3568..3636))
                     /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     3399..3418
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3637..3729)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3810..4398
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4299..4318
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4552..4569
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4584..4726
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4652..4671)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4812..4834
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4828..5019)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5591..6673)
                     /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     6635..6654
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6674..6751)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6957..6976
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(7064..7080,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(7064..7080,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac tttaagaagg
       61 agatatcata tgggtggtgg tagcggtggt tctggtggtt ctggtggtag cggcggtagc
      121 ggtggtagcg gtggtagcgg tggcagcggc ggttctagca aaggtgaacg tctgtttcgt
      181 ggtaaagtac cgatcttagt ggaattaaag ggcgacgtga acggtcataa atttagcgtg
      241 cgcggcaaag gcaaaggtga cgctacccgt ggtaaattga ccctgaagtt tatttgcaca
      301 acaggcaaat tacccgttcc gtggcccacc ttagtgacca ccctgaccta tggcgttcag
      361 tgcttcagtc gttaccctaa acacatgaaa cgtcacgatt ttttcaaatc agccatgcct
      421 aaaggatatg ttcaagagcg tacaatcagc ttcaagaagg atggcaaata taaaacgcgt
      481 gcggaagtga aatttgaagg ccgcacatta gtaaatcgta tcaaactgaa aggtcgtgac
      541 ttcaaagaaa aaggcaacat tttaggccat aaactgcgtt ataactttaa ttctcataag
      601 gtgtatatta cggccgataa acgcaagaat ggtatcaagg caaaattcaa aattcgccat
      661 aacgtgaaag acggcagcgt tcaattagcg gatcattatc aacaaaacac gccgattggt
      721 cgcgggcctg tactgttacc tcgcaaccac tacctgagca cccgttctaa actgagcaaa
      781 gatccgaaag aaaaacgcga tcacatggtt ctgttagaat tcgtgaccgc tgcaggcatt
      841 aagcacggac gcgacgaacg ctacaagacc ggtggtagcg gtggttctgg tggttctggt
      901 ggtagcggcg gtagcggtgg tagcggtggt agcggtggca gcggcggtac cgcgagcaat
      961 ttactgaccg tacaccaaaa tttgcctgca ttgccggtcg atgcaacgag tgatgaggtt
     1021 cgcaagaacc tgatggacat gttcagggat cgccaggcgt tttctgagca tacctggaaa
     1081 atgcttctgt ccgtttgccg gtcgtgggcg gcatggtgca agttgaataa ccggaaatgg
     1141 tttcccgcag aacctgaaga tgttcgcgat tatcttctat atcttcaggc gcgcggtctg
     1201 gcagtaaaaa ctatccagca acatttgggc cagctaaaca tgcttcatcg tcggtccggg
     1261 ctgccacgac caagtgacag caatgctgtt tcactggtta tgcggcgtat ccgaaaagaa
     1321 aacgttgatg ccggtgaacg tgcaaaacag gctctagcgt tcgaacgcac tgatttcgac
     1381 caggttcgtt cactcatgga aaatagcgat cgctgccagg atatacgtaa tctggcattt
     1441 ctggggattg cttataacac cctgttacgt atagccgaaa ttgccaggat cagggttaaa
     1501 gatatctcac gtactgacgg tgggagaatg ttaatccata ttggcagaac gaaaacgctg
     1561 gttagcaccg caggtgtaga gaaggcactt agcctggggg taactaaact ggtcgagcga
     1621 tggatttccg tctctggtgt agctgatgat ccgaataact acctgttttg ccgggtcaga
     1681 aaaaatggtg ttgccgcgcc atctgccacc agccagctat caactcgcgc cctggaaggg
     1741 atttttgaag caactcatcg attgatttac ggcgctaagg atgactctgg tcagagatac
     1801 ctggcctggt ctggacacag tgcccgtgtc ggagccgcgc gagatatggc ccgcgctgga
     1861 gtttcaatac cggagatcat gcaagctggt ggctggacca atgtaaatat tgtcatgaac
     1921 tatatccgta acctggatag tgaaacaggg gcaatggtgc gcctgctgga agatggcgac
     1981 ggcggatccc atcaccacca ccatcactaa tgactcgagt ctggtaaaga aaccgctgct
     2041 gcgaaatttg aacgccagca catggactcg tctactagcg cagcttaatt aacctaggct
     2101 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     2161 ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg ccctgtagcg
     2221 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     2281 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     2341 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     2401 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     2461 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     2521 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     2581 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     2641 aaatattaac gtttacaatt tctggcggca cgatggcatg agattatcaa aaaggatctt
     2701 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     2761 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     2821 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     2881 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     2941 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     3001 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3061 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3121 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     3181 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     3241 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     3301 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     3361 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     3421 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     3481 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     3541 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     3601 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatcatgatt
     3661 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     3721 ataaacaaat aggtcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     3781 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     3841 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     3901 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     3961 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     4021 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     4081 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     4141 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     4201 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     4261 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     4321 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     4381 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     4441 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     4501 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     4561 gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     4621 atttcacacc gcatatatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     4681 gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc
     4741 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     4801 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     4861 gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg
     4921 ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa
     4981 gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg
     5041 atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt
     5101 tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg
     5161 gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga
     5221 tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt
     5281 gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca
     5341 tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat
     5401 cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga
     5461 caggagcacg atcatgctag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa
     5521 ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat
     5581 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     5641 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt
     5701 tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca
     5761 gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg
     5821 gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag atgtccgcac
     5881 caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg
     5941 caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac
     6001 cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga
     6061 gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta
     6121 acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt
     6181 cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg
     6241 ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt
     6301 taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt
     6361 cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag
     6421 atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc
     6481 caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca
     6541 gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt
     6601 tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg
     6661 ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac
     6721 cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac
     6781 tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg
     6841 aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata
     6901 cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg
     6961 atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg
     7021 cgtccggcgt agaggatcga gatcgatctc gatcccgcga aattaatacg actcactata
//
