LOCUS       sequence-135604-        7149 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  His-TEV-U+36GFP-Cre for E.Coli expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7149)
  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 7149)
  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: His-TEV-U_(+36GFP)-Cre
FEATURES             Location/Qualifiers
     source          1..7149
                     /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             73..90
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             91..111
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1042..2067
                     /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             2095..2139
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2177..2195)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2191..2238
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2275..2730
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2362..2381)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2572..2593
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2845..3636,3637..3705))
                     /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     3468..3487
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3706..3798)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3879..4467
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4368..4387
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4621..4638
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4653..4795
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4721..4740)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4881..4903
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4897..5088)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5660..6742)
                     /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     6704..6723
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6743..6820)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7026..7045
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(7133..7149,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(7133..7149,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac tttaagaagg
       61 agatatcata tgcaccatca ccatcaccat gaaaacctgt acttccaggg cctgtttgat
      121 attattaaaa aagtggcgag cgtgattggc ggcctggcta gcggtggtag cggtggttct
      181 ggtggttctg gtggtagcgg cggtagcggt ggtagcggtg gtagcggtgg cagcggcggt
      241 tctagcaaag gtgaacgtct gtttcgtggt aaagtaccga tcttagtgga attaaagggc
      301 gacgtgaacg gtcataaatt tagcgtgcgc ggcaaaggca aaggtgacgc tacccgtggt
      361 aaattgaccc tgaagtttat ttgcacaaca ggcaaattac ccgttccgtg gcccacctta
      421 gtgaccaccc tgacctatgg cgttcagtgc ttcagtcgtt accctaaaca catgaaacgt
      481 cacgattttt tcaaatcagc catgcctaaa ggatatgttc aagagcgtac aatcagcttc
      541 aagaaggatg gcaaatataa aacgcgtgcg gaagtgaaat ttgaaggccg cacattagta
      601 aatcgtatca aactgaaagg tcgtgacttc aaagaaaaag gcaacatttt aggccataaa
      661 ctgcgttata actttaattc tcataaggtg tatattacgg ccgataaacg caagaatggt
      721 atcaaggcaa aattcaaaat tcgccataac gtgaaagacg gcagcgttca attagcggat
      781 cattatcaac aaaacacgcc gattggtcgc gggcctgtac tgttacctcg caaccactac
      841 ctgagcaccc gttctaaact gagcaaagat ccgaaagaaa aacgcgatca catggttctg
      901 ttagaattcg tgaccgctgc aggcattaag cacggacgcg acgaacgcta caagaccggt
      961 ggtagcggtg gttctggtgg ttctggtggt agcggcggta gcggtggtag cggtggtagc
     1021 ggtggcagcg gcggtaccgc gagcaattta ctgaccgtac accaaaattt gcctgcattg
     1081 ccggtcgatg caacgagtga tgaggttcgc aagaacctga tggacatgtt cagggatcgc
     1141 caggcgtttt ctgagcatac ctggaaaatg cttctgtccg tttgccggtc gtgggcggca
     1201 tggtgcaagt tgaataaccg gaaatggttt cccgcagaac ctgaagatgt tcgcgattat
     1261 cttctatatc ttcaggcgcg cggtctggca gtaaaaacta tccagcaaca tttgggccag
     1321 ctaaacatgc ttcatcgtcg gtccgggctg ccacgaccaa gtgacagcaa tgctgtttca
     1381 ctggttatgc ggcgtatccg aaaagaaaac gttgatgccg gtgaacgtgc aaaacaggct
     1441 ctagcgttcg aacgcactga tttcgaccag gttcgttcac tcatggaaaa tagcgatcgc
     1501 tgccaggata tacgtaatct ggcatttctg gggattgctt ataacaccct gttacgtata
     1561 gccgaaattg ccaggatcag ggttaaagat atctcacgta ctgacggtgg gagaatgtta
     1621 atccatattg gcagaacgaa aacgctggtt agcactgcag gtgtagagaa ggcacttagc
     1681 ctgggggtaa ctaaactggt cgagcgatgg atttccgtct ctggtgtagc tgatgatccg
     1741 aataactacc tgttttgccg ggtcagaaaa aatggtgttg ccgcgccatc tgccaccagc
     1801 cagctatcaa ctcgcgccct ggaagggatt tttgaagcaa ctcatcgatt gatttacggc
     1861 gctaaggatg actctggtca gagatacctg gcctggtctg gacacagtgc ccgtgtcgga
     1921 gccgcgcgag atatggcccg cgctggagtt tcaataccgg agatcatgca agctggtggc
     1981 tggaccaatg taaatattgt catgaactat atccgtaacc tggatagtga aacaggggca
     2041 atggtgcgcc tgctggaaga tggcgacggc ggatcctaat gactcgagtc tggtaaagaa
     2101 accgctgctg cgaaatttga acgccagcac atggactcgt ctactagcgc agcttaatta
     2161 acctaggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg
     2221 gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa tgggacgcgc
     2281 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     2341 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     2401 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     2461 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     2521 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     2581 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     2641 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     2701 attttaacaa aatattaacg tttacaattt ctggcggcac gatggcatga gattatcaaa
     2761 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     2821 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     2881 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     2941 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     3001 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     3061 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     3121 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     3181 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     3241 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     3301 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     3361 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     3421 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     3481 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     3541 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     3601 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     3661 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     3721 atcatgattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     3781 tttagaaaaa taaacaaata ggtcatgacc aaaatccctt aacgtgagtt ttcgttccac
     3841 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     3901 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     3961 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     4021 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     4081 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     4141 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     4201 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     4261 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     4321 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     4381 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     4441 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     4501 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     4561 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     4621 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     4681 ctgtgcggta tttcacaccg catatatggt gcactctcag tacaatctgc tctgatgccg
     4741 catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg
     4801 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     4861 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     4921 gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat
     4981 gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct
     5041 tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt
     5101 gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac
     5161 gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact
     5221 ggcggtatgg atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt
     5281 taatacagat gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa
     5341 cataatggtg cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa
     5401 gaccattcat gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg
     5461 ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt
     5521 cctcaacgac aggagcacga tcatgctagt catgccccgc gcccaccgga aggagctgac
     5581 tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag tgagctaact
     5641 tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     5701 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gccagggtgg
     5761 tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc tggccctgag
     5821 agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc tgtttgatgg
     5881 tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc actaccgaga
     5941 tgtccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc agcgccatct
     6001 gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt tgcatggttt
     6061 gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc tgaatttgat
     6121 tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca gaacttaatg
     6181 ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc acgcccagtc
     6241 gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca gagacatcaa
     6301 gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc tggtcatcca
     6361 gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc accgccgctt
     6421 tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca cccagttgat
     6481 cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc agactggagg
     6541 tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg cggttgggaa
     6601 tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca gaaacgtggc
     6661 tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac tctgcgacat
     6721 cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc gggcgctatc
     6781 atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg acgctctccc
     6841 ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt gagcaccgcc
     6901 gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc cacggggcct
     6961 gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc ccgatcttcc
     7021 ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg
     7081 gccacgatgc gtccggcgta gaggatcgag atcgatctcg atcccgcgaa attaatacga
     7141 ctcactata
//
