LOCUS       sequence-464125-        6029 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of epsin-1 (EPS-15-interacting 
            protein 1) in Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6029)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 6029)
  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"
COMMENT     Sequence Label: pAIP4_1EYH
FEATURES             Location/Qualifiers
     source          1..6029
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(126..173)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     169..187
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(240..257)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1014..1043)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             1066..1088
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1103..1127
                     /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)."
     primer_bind     complement(1127..1146)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1128..1146)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1234..1253)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1459..1536
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1537..2619
                     /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     complement(1556..1575)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3428..3619
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3613..3635)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3721..3860
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3776..3795
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3875..3892)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4046..4634)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4126..4145)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4745..4764)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4756..5571
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4756..5571
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4829..4848)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(5663..6029,1..89))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5800..5821)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     join(6012..6029,1..2)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
       61 cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca atccggatat agttcctcct
      121 ttcagcaaaa aacccctcaa gacccgttta gaggccccaa ggggttatgc tagttattgc
      181 tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt tgttagcagc cggatctcag
      241 tggtggtggt ggtggtgctc gagtcattac gaacctgcgg tttgagccag cttttctttg
      301 gtcttgagcg catgtgcgcg ttcctcgcgc aaacgatctt cgtcacgcag caaggcaacc
      361 agctgtttcg ccttctcacg aacattaacg ccctggtcct taccatcacg atccacgtat
      421 tggaagtcct taagagtttg aaccgcgtac atgttctctt tgcactgctg gctcacgcgc
      481 tcgctacccg ttttaatcag atattccatc agggtcatag ctttgtacac gtgacgccag
      541 ttcttgccgt ggtcattcaa acgtttccaa atcatggaca tgatctcgct aaacgccacg
      601 acgttatagg tcagatcggc gatttcggac atcagagacg agctcggacc ccacgggtcg
      661 ttgctggtcg cttctcttac ttttatttca gcctctgaat aattgtgtcc agagccgcca
      721 atctgttcac ggtgggcttc aatgatgtcg ttgtcttcca tgtccaggtc ttccggcgct
      781 tgatctgcct ggatgcgaat accgtcatac aggaaacgca ggctatccat ttccttgccc
      841 tgacgtttgg caaacgcttc catcaggcga cgcagcggcg tggtcttttt gatcttaaag
      901 aaaatttcag agctgccgtc actaactttc aggttaatat gggtttccgg tttcacttcc
      961 ggcttgactt ccggctttgc ttcttggttg acttccgaat cttgcaggct accgtggtga
     1021 tggtgatggt gatgatggtg atgaatcttc gccatcatat gtatatctcc ttcttaaagt
     1081 taaacaaaat tatttctaga ggggaattgt tatccgctca caattcccct atagtgagtc
     1141 gtattaattt cgcgggatcg agatcgatct cgatcctcta cgccggacgc atcgtggccg
     1201 gcatcaccgg cgccacaggt gcggttgctg gcgcctatat cgccgacatc accgatgggg
     1261 aagatcgggc tcgccacttc gggctcatga gcgcttgttt cggcgtgggt atggtggcag
     1321 gccccgtggc cgggggactg ttgggcgcca tctccttgca tgcaccattc cttgcggcgg
     1381 cggtgctcaa cggcctcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg
     1441 gagagcgtcg agatcccgga caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga
     1501 tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat
     1561 gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc
     1621 cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt
     1681 cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc
     1741 tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat
     1801 caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa
     1861 gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg
     1921 gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt
     1981 gatgtctctg accagacacc catcaacagt attattttct cccatgaaga cggtacgcga
     2041 ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca
     2101 ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat
     2161 caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa
     2221 accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag
     2281 atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggacatc
     2341 tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc gttaaccacc
     2401 atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct gcaactctct
     2461 cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa aagaaaaacc
     2521 accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     2581 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag
     2641 ttagctcact cattaggcac cgggatctcg accgatgccc ttgagagcct tcaacccagt
     2701 cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt
     2761 tatcatgcaa ctcgtaggac aggtgccggc agcgctctgg gtcattttcg gcgaggaccg
     2821 ctttcgctgg agcgcgacga tgatcggcct gtcgcttgcg gtattcggaa tcttgcacgc
     2881 cctcgctcaa gccttcgtca ctggtcccgc caccaaacgt ttcggcgaga agcaggccat
     2941 tatcgccggc atggcggccc cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg
     3001 gctaggctgg cggggttgcc ttactggtta gcagaatgaa tcaccgatac gcgagcgaac
     3061 gtgaagcgac tgctgctgca aaacgtctgc gacctgagca acaacatgaa tggtcttcgg
     3121 tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg
     3181 gatctgcatc gcaggatgct gctggctacc ctgtggaaca cctacatctg tattaacgaa
     3241 gcgctggcat tgaccctgag tgatttttct ctggtcccgc cgcatccata ccgccagttg
     3301 tttaccctca caacgttcca gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag
     3361 catcctctct cgtttcatcg gtatcattac ccccatgaac agaaatcccc cttacacgga
     3421 ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca
     3481 gacattaacg cttctggaga aactcaacga gctggacgcg gatgaacagg cagacatctg
     3541 tgaatcgctt cacgaccacg ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga
     3601 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     3661 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
     3721 cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca
     3781 tcagagcaga ttgtactgag agtgcaccat tgcggtgtga aataccgcac agatgcgtaa
     3841 ggagaaaata ccgcatcagg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3901 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3961 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4021 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4081 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4141 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4201 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4261 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4321 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4381 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4441 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     4501 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4561 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4621 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4681 aaaactcacg ttaagggatt ttggtcatga acaataaaac tgtctgctta cataaacagt
     4741 aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cttgctctag gccgcgatta
     4801 aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa
     4861 tcaggtgcga caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa
     4921 catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg
     4981 acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg
     5041 ttactcacca ctgcgatccc cgggaaaaca gcattccagg tattagaaga atatcctgat
     5101 tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct
     5161 gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga
     5221 atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt
     5281 gaacaagtct ggaaagaaat gcataaactt ttgccattct caccggattc agtcgtcact
     5341 catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt
     5401 gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc
     5461 ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat
     5521 cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta agaattaatt
     5581 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     5641 atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa attcgcgtta
     5701 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     5761 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     5821 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     5881 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     5941 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     6001 gcgagaaagg aagggaagaa agcgaaagg
//
