LOCUS       sequence-133162-        7131 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  U+36GFP-Cre fusion expression in E. Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7131)
  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 7131)
  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: U_(+36GFP)-Cre.ape
FEATURES             Location/Qualifiers
     source          1..7131
                     /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             1006..2031
                     /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             2041..2058
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2077..2121
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2159..2177)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2173..2220
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2257..2712
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2344..2363)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2554..2575
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2827..3618,3619..3687))
                     /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     3450..3469
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3688..3780)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3861..4449
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4350..4369
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4603..4620
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4635..4777
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4703..4722)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4863..4885
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4879..5070)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5642..6724)
                     /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     6686..6705
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6725..6802)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7008..7027
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(7115..7131,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(7115..7131,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac tttaagaagg
       61 agatatcata tgggcctgtt tgatattatt aaaaaagtgg cgagcgtgat tggcggcctg
      121 gctagcggtg gtagcggtgg ttctggtggt tctggtggta gcggcggtag cggtggtagc
      181 ggtggtagcg gtggcagcgg cggttctagc aaaggtgaac gtctgtttcg tggtaaagta
      241 ccgatcttag tggaattaaa gggcgacgtg aacggtcata aatttagcgt gcgcggcaaa
      301 ggcaaaggtg acgctacccg tggtaaattg accctgaagt ttatttgcac aacaggcaaa
      361 ttacccgttc cgtggcccac cttagtgacc accctgacct atggcgttca gtgcttcagt
      421 cgttacccta aacacatgaa acgtcacgat tttttcaaat cagccatgcc taaaggatat
      481 gttcaagagc gtacaatcag cttcaagaag gatggcaaat ataaaacgcg tgcggaagtg
      541 aaatttgaag gccgcacatt agtaaatcgt atcaaactga aaggtcgtga cttcaaagaa
      601 aaaggcaaca ttttaggcca taaactgcgt tataacttta attctcataa ggtgtatatt
      661 acggccgata aacgcaagaa tggtatcaag gcaaaattca aaattcgcca taacgtgaaa
      721 gacggcagcg ttcaattagc ggatcattat caacaaaaca cgccgattgg tcgcgggcct
      781 gtactgttac ctcgcaacca ctacctgagc acccgttcta aactgagcaa agatccgaaa
      841 gaaaaacgcg atcacatggt tctgttagaa ttcgtgaccg ctgcaggcat taagcacgga
      901 cgcgacgaac gctacaagac cggtggtagc ggtggttctg gtggttctgg tggtagcggc
      961 ggtagcggtg gtagcggtgg tagcggtggc agcggcggta ccgcgagcaa tttactgacc
     1021 gtacaccaaa atttgcctgc attgccggtc gatgcaacga gtgatgaggt tcgcaagaac
     1081 ctgatggaca tgttcaggga tcgccaggcg ttttctgagc atacctggaa aatgcttctg
     1141 tccgtttgcc ggtcgtgggc ggcatggtgc aagttgaata accggaaatg gtttcccgca
     1201 gaacctgaag atgttcgcga ttatcttcta tatcttcagg cgcgcggtct ggcagtaaaa
     1261 actatccagc aacatttggg ccagctaaac atgcttcatc gtcggtccgg gctgccacga
     1321 ccaagtgaca gcaatgctgt ttcactggtt atgcggcgta tccgaaaaga aaacgttgat
     1381 gccggtgaac gtgcaaaaca ggctctagcg ttcgaacgca ctgatttcga ccaggttcgt
     1441 tcactcatgg aaaatagcga tcgctgccag gatatacgta atctggcatt tctggggatt
     1501 gcttataaca ccctgttacg tatagccgaa attgccagga tcagggttaa agatatctca
     1561 cgtactgacg gtgggagaat gttaatccat attggcagaa cgaaaacgct ggttagcact
     1621 gcaggtgtag agaaggcact tagcctgggg gtaactaaac tggtcgagcg atggatttcc
     1681 gtctctggtg tagctgatga tccgaataac tacctgtttt gccgggtcag aaaaaatggt
     1741 gttgccgcgc catctgccac cagccagcta tcaactcgcg ccctggaagg gatttttgaa
     1801 gcaactcatc gattgattta cggcgctaag gatgactctg gtcagagata cctggcctgg
     1861 tctggacaca gtgcccgtgt cggagccgcg cgagatatgg cccgcgctgg agtttcaata
     1921 ccggagatca tgcaagctgg tggctggacc aatgtaaata ttgtcatgaa ctatatccgt
     1981 aacctggata gtgaaacagg ggcaatggtg cgcctgctgg aagatggcga cggcggatcc
     2041 catcaccacc accatcacta atgactcgag tctggtaaag aaaccgctgc tgcgaaattt
     2101 gaacgccagc acatggactc gtctactagc gcagcttaat taacctaggc tgctgccacc
     2161 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     2221 ctgaaaggag gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa
     2281 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     2341 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     2401 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     2461 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     2521 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     2581 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     2641 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     2701 cgtttacaat ttctggcggc acgatggcat gagattatca aaaaggatct tcacctagat
     2761 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     2821 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     2881 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     2941 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     3001 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     3061 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     3121 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     3181 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     3241 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     3301 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     3361 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     3421 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     3481 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     3541 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     3601 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     3661 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatcatgat tgaagcattt
     3721 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     3781 taggtcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     3841 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3901 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3961 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     4021 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     4081 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     4141 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     4201 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     4261 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     4321 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     4381 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     4441 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     4501 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     4561 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     4621 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     4681 cgcatatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtata
     4741 cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc
     4801 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     4861 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagct
     4921 gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct gttcatccgc
     4981 gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa agcgggccat
     5041 gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt
     5101 catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg ttactgatga
     5161 tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg
     5221 ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag atgtaggtgt
     5281 tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg tgcagggcgc
     5341 tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc atgttgttgc
     5401 tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc
     5461 attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg acaggagcac
     5521 gatcatgcta gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa
     5581 gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg
     5641 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5701 acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg
     5761 agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt
     5821 ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat
     5881 aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatgtccgca ccaacgcgca
     5941 gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca
     6001 tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg
     6061 cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat
     6121 gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga
     6181 tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg
     6241 agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat
     6301 tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca
     6361 gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc
     6421 ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg
     6481 ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca
     6541 acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca
     6601 tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc
     6661 gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt
     6721 tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg
     6781 ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt
     6841 aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca
     6901 tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg
     6961 aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
     7021 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
     7081 tagaggatcg agatcgatct cgatcccgcg aaattaatac gactcactat a
//
