LOCUS       sequence-133188-        7053 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Aurein1.2+36GFP-BirA fusion for E.Coli expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7053)
  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 7053)
  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: Aurein1.2_(+36GFP)-BirA
FEATURES             Location/Qualifiers
     source          1..7053
                     /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             994..1953
                     /codon_start=1
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             994..1953
                     /codon_start=1
                     /gene="birA"
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             1963..1980
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1999..2043
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2081..2099)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2095..2142
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2179..2634
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2266..2285)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2476..2497
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2749..3540,3541..3609))
                     /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     3372..3391
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3610..3702)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3783..4371
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4272..4291
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4525..4542
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4557..4699
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4625..4644)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4785..4807
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4801..4992)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5564..6646)
                     /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     6608..6627
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6647..6724)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6930..6949
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(7037..7053,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(7037..7053,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac tttaagaagg
       61 agatatcata tgggcctgtt tgatattatt aaaaaaattg cggaaagctt tgctagcggt
      121 ggtagcggtg gttctggtgg ttctggtggt agcggcggta gcggtggtag cggtggtagc
      181 ggtggcagcg gcggttctag caaaggtgaa cgtctgtttc gtggtaaagt accgatctta
      241 gtggaattaa agggcgacgt gaacggtcat aaatttagcg tgcgcggcaa aggcaaaggt
      301 gacgctaccc gtggtaaatt gaccctgaag tttatttgca caacaggcaa attacccgtt
      361 ccgtggccca ccttagtgac caccctgacc tatggcgttc agtgcttcag tcgttaccct
      421 aaacacatga aacgtcacga ttttttcaaa tcagccatgc ctaaaggata tgttcaagag
      481 cgtacaatca gcttcaagaa ggatggcaaa tataaaacgc gtgcggaagt gaaatttgaa
      541 ggccgcacat tagtaaatcg tatcaaactg aaaggtcgtg acttcaaaga aaaaggcaac
      601 attttaggcc ataaactgcg ttataacttt aattctcata aggtgtatat tacggccgat
      661 aaacgcaaga atggtatcaa ggcaaaattc aaaattcgcc ataacgtgaa agacggcagc
      721 gttcaattag cggatcatta tcaacaaaac acgccgattg gtcgcgggcc tgtactgtta
      781 cctcgcaacc actacctgag cacccgttct aaactgagca aagatccgaa agaaaaacgc
      841 gatcacatgg ttctgttaga attcgtgacc gctgcaggca ttaagcacgg acgcgacgaa
      901 cgctacaaga ccggtggttc aggcggtagt ggtggtagtg gcggttcagg tggtagtggt
      961 ggctcaggtg gcagtggtgg ttcaggtggt agtaaggata acaccgtgcc actgaaattg
     1021 attgccctgt tagcgaacgg tgaatttcac tctggcgagc agttgggtga aacgctggga
     1081 atgagccggg cggctattaa taaacacatt cagacactgc gtgactgggg cgttgatgtc
     1141 tttaccgttc cgggtaaagg atacagcctg cctgagccta tccagttact taatgctaaa
     1201 cagatattgg gtcagctgga tggcggtagt gtagccgtgc tgccagtgat tgactccacg
     1261 aatcagtacc ttcttgatcg tatcggagag cttaaatcgg gcgatgcttg cattgcagaa
     1321 taccagcagg ctggccgtgg tcgccggggt cggaaatggt tttcgccttt tggcgcaaac
     1381 ttatatttgt cgatgttctg gcgtctggaa caaggcccgg cggcggcgat tggtttaagt
     1441 ctggttatcg gtatcgtgat ggcggaagta ttacgcaagc tgggtgcaga taaagttcgt
     1501 gttaaatggc ctaatgacct ctatctgcag gatcgcaagc tggcaggcat tctggttgag
     1561 ctgactggca aaactggcga tgcggcgcaa atagtcattg gagccgggat caacatggca
     1621 atgcgccgtg ttgaagagag tgtcgttaat caggggtgga tcacgctgca ggaagcgggg
     1681 atcaatctcg atcgtaatac gttggcggcc atgctaatac gtgaattacg tgctgcgttg
     1741 gaactcttcg aacaagaagg attggcacct tatctgtcgc gctgggaaaa gctggataat
     1801 tttattaatc gcccagtgaa acttatcatt ggtgataaag aaatatttgg catttcacgc
     1861 ggaatagaca aacagggggc tttattactt gagcaggatg gaataataaa accctggatg
     1921 ggcggtgaaa tatccctgcg tagtgcagaa aaaaggggat cccatcacca ccaccatcac
     1981 taatgactcg agtctggtaa agaaaccgct gctgcgaaat ttgaacgcca gcacatggac
     2041 tcgtctacta gcgcagctta attaacctag gctgctgcca ccgctgagca ataactagca
     2101 taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata
     2161 tccggattgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     2221 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     2281 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     2341 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     2401 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     2461 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     2521 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     2581 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca atttctggcg
     2641 gcacgatggc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     2701 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     2761 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     2821 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     2881 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     2941 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     3001 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     3061 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     3121 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     3181 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     3241 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     3301 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     3361 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     3421 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     3481 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     3541 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     3601 aatactcata ctcttccttt ttcaatcatg attgaagcat ttatcagggt tattgtctca
     3661 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggtcat gaccaaaatc
     3721 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     3781 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     3841 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     3901 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     3961 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     4021 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     4081 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     4141 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     4201 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     4261 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     4321 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     4381 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     4441 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     4501 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg
     4561 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatata tggtgcactc
     4621 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg
     4681 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     4741 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     4801 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
     4861 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
     4921 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
     4981 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
     5041 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
     5101 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
     5161 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
     5221 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
     5281 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt
     5341 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
     5401 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc tagtcatgcc
     5461 ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc
     5521 cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca
     5581 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     5641 tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat
     5701 tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca
     5761 gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg
     5821 tatcgtcgta tcccactacc gagatgtccg caccaacgcg cagcccggac tcggtaatgg
     5881 cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc
     5941 cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc
     6001 gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca
     6061 gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg
     6121 cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga
     6181 tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca
     6241 cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg
     6301 cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca
     6361 ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg
     6421 gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca
     6481 gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt
     6541 cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag
     6601 agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc accaccctga
     6661 attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg
     6721 tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt
     6781 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc
     6841 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     6901 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     6961 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cgagatcgat
     7021 ctcgatcccg cgaaattaat acgactcact ata
//
