LOCUS       sequence-130233-        6063 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  His-TEV-Aurein1.2+36GFP-LPETG-His expression in E.Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6063)
  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 6063)
  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-Aurein1.2_(+36GFP)-LPETG-His
FEATURES             Location/Qualifiers
     source          1..6063
                     /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             973..990
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1009..1053
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1091..1109)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1105..1152
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1189..1644
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1276..1295)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1486..1507
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(1759..2550,2551..2619))
                     /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     2382..2401
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2620..2712)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2793..3381
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3282..3301
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3535..3552
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3567..3709
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3635..3654)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3795..3817
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3811..4002)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4574..5656)
                     /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     5618..5637
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5657..5734)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5940..5959
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(6047..6063,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(6047..6063,1..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 aaattgcgga aagctttgct agcggtggta gcggtggttc tggtggttct
      181 ggtggtagcg gcggtagcgg tggtagcggt ggtagcggtg gcagcggcgg ttctagcaaa
      241 ggtgaacgtc tgtttcgtgg taaagtaccg atcttagtgg aattaaaggg cgacgtgaac
      301 ggtcataaat ttagcgtgcg cggcaaaggc aaaggtgacg ctacccgtgg taaattgacc
      361 ctgaagttta tttgcacaac aggcaaatta cccgttccgt ggcccacctt agtgaccacc
      421 ctgacctatg gcgttcagtg cttcagtcgt taccctaaac acatgaaacg tcacgatttt
      481 ttcaaatcag ccatgcctaa aggatatgtt caagagcgta caatcagctt caagaaggat
      541 ggcaaatata aaacgcgtgc ggaagtgaaa tttgaaggcc gcacattagt aaatcgtatc
      601 aaactgaaag gtcgtgactt caaagaaaaa ggcaacattt taggccataa actgcgttat
      661 aactttaatt ctcataaggt gtatattacg gccgataaac gcaagaatgg tatcaaggca
      721 aaattcaaaa ttcgccataa cgtgaaagac ggcagcgttc aattagcgga tcattatcaa
      781 caaaacacgc cgattggtcg cgggcctgta ctgttacctc gcaaccacta cctgagcacc
      841 cgttctaaac tgagcaaaga tccgaaagaa aaacgcgatc acatggttct gttagaattc
      901 gtgaccgctg caggcattaa gcacggacgc gacgaacgct acaagaccgg tggtagcctc
      961 ccggaaaccg gtcatcacca ccaccatcac taatgactcg agtctggtaa agaaaccgct
     1021 gctgcgaaat ttgaacgcca gcacatggac tcgtctacta gcgcagctta attaacctag
     1081 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
     1141 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
     1201 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     1261 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     1321 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     1381 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     1441 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     1501 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     1561 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     1621 acaaaatatt aacgtttaca atttctggcg gcacgatggc atgagattat caaaaaggat
     1681 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     1741 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     1801 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     1861 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     1921 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     1981 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     2041 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     2101 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     2161 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     2221 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     2281 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     2341 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     2401 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     2461 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     2521 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     2581 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatcatg
     2641 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     2701 aaaataaaca aataggtcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     2761 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     2821 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     2881 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     2941 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3001 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3061 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     3121 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     3181 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     3241 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     3301 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     3361 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     3421 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     3481 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     3541 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     3601 ggtatttcac accgcatata tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     3661 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     3721 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     3781 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     3841 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc
     3901 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat
     3961 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg
     4021 gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg
     4081 ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt
     4141 atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac
     4201 agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat
     4261 ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat
     4321 tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg
     4381 tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa
     4441 cgacaggagc acgatcatgc tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt
     4501 gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct aacttacatt
     4561 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4621 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc
     4681 ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt
     4741 gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta
     4801 acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc gagatgtccg
     4861 caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt
     4921 tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa
     4981 aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag
     5041 tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg
     5101 ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac
     5161 cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata
     5221 acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat
     5281 agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg
     5341 cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc
     5401 gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa
     5461 cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat
     5521 tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct
     5581 ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata
     5641 acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca
     5701 taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc tcccttatgc
     5761 gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca
     5821 aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc
     5881 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
     5941 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
     6001 atgcgtccgg cgtagaggat cgagatcgat ctcgatcccg cgaaattaat acgactcact
     6061 ata
//
