LOCUS       sequence-482865-        6609 bp    DNA     circular SYN 11-AUG-2025
DEFINITION  Express IL13Ra1-HaloTag Fusion Protein in Mammalian Cells under a 
            CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6609)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6609)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 11, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: IL13Ra1-HaloTag BiBRET Vector
FEATURES             Location/Qualifiers
     source          1..6609
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..546
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     447..466
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     663..680
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    830..878
                     /note="synthetic polyadenylation signal"
     CDS             complement(902..1300)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      1297..1306
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(1331..1688)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1345..1480
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        1497..1688
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     1645..1665
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    1767..1848
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     enhancer        2081..2384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2385..2588
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2539..2559
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          2724..2856
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2839..2858
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     2900..2919
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2900..2918
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4234..4254
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             4270..5154
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     polyA_signal    5241..5362
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5252..5271)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(5477..5510)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5520..5567
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5599..6393
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5653..5672)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6263..6282
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 ttgagatcgt tttggtctgc gcgtaatctc ttgctctgaa aacgaaaaaa ccgccttgca
       61 gggcggtttt tcgaaggttc tctgagctac caactctttg aaccgaggta actggcttgg
      121 aggagcgcag tcaccaaaac ttgtcctttc agtttagcct taaccggcgc atgacttcaa
      181 gactaactcc tctaaatcaa ttaccagtgg ctgctgccag tggtgctttt gcatgtcttt
      241 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcggac tgaacggggg
      301 gttcgtgcat acagtccagc ttggagcgaa ctgcctaccc ggaactgagt gtcaggcgtg
      361 gaatgagaca aacgcggcca taacagcgga atgacaccgg taaaccgaaa ggcaggaaca
      421 ggagagcgca cgagggagcc gccaggggga aacgcctggt atctttatag tcctgtcggg
      481 tttcgccacc actgatttga gcgtcagatt tcgtgatgct tgtcaggggg gcggagccta
      541 tggaaaaacg gctttgccgc ggccctctca cttccctgtt aagtatcttc ctggcatctt
      601 ccaggaaatc tccgccccgt tcgtaagcca tttccgctcg ccgcagtcga acgaccgagc
      661 gtagcgagtc agtgagcgag gaagcggaat atatcctgta tcacatattc tgctgacgca
      721 ccggtgcagc cttttttctc ctgccacatg aagcacttca ctgacaccct catcagtgcc
      781 aacatagtaa gccagtatac actccgctag cgctgatgtc cggaatcgtc acacaaaaaa
      841 ccaacacaca gatgtaatga aaataaagat attttattgc ggccatcgtg atggctagag
      901 tttagccctc ccacacataa ccagagggca gcaattcacg aatcccaact gccgtcggct
      961 gtccatcact gtccttcact atggctttga tcccaggatg cagatcgaga agcacctgtc
     1021 ggcaccgtcc gcaggggctc aagatgcccc tgttctcatt tccgatcgcg acgatacaag
     1081 tcaggttgcc agctgccgca gcagcagcag tgcccagcac cacgagttct gcacaaggtc
     1141 ccccagtaaa atgatataca ttgacaccag tgaagatgcg gccgtcgcta gagagagctg
     1201 cgctggcgac gctgtagtct tcagagatgg ggatgctgtt gattgtagcc gttgctcttt
     1261 caatgagggt ggattcttct tgagacaaag gcttggccat ggtggcgcta gtgtcagaag
     1321 aatcgagctt tttgcaaaag cctaggcctc caaaaaagcc tcctcactac ttctggaata
     1381 gctcagaggc cgaggcggcc tcggcctctg cataaataaa aaaaattagt cagccatggg
     1441 gcggagaatg ggcggaactg ggcggagtta ggggcgggat gggcggagtt aggggcggga
     1501 ctatggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg
     1561 gggactttcc acacctggtt gctgactaat tgagatgcat gctttgcata cttctgcctg
     1621 ctggggagcc tggggacttt ccacacccta actgacacac attccacagc tggttctttc
     1681 cgcctcagaa ggtacctaac caagttcctc tttcagaggt tatttcaggc catggtgctg
     1741 cgcaagacgc gttaattaag acgtcggcag tgaaaaaaat gctttatttg tgaaatttgt
     1801 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     1861 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa
     1921 aacctctaca aatgtggtat ggctgattat gatccgcggc cgcaatacgt cgacgttatc
     1981 agctgacttc gtacgagagc ctaggattat ggcgcgccac tagttattaa tagtaatcaa
     2041 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
     2101 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
     2161 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
     2221 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtccgccc cctattgacg
     2281 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta cgggactttc
     2341 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
     2401 agtacaccaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
     2461 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
     2521 ataaccccgc cccgttgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
     2581 gcagagctgg tttagtgaac cgtcagatca ctagaagctt tattgcggta gtttatcaca
     2641 gttaaattgc taacgcagtc agtgcttctg acacaacagt ctcgaactta agctgcagaa
     2701 gttggtcgtg aggcactggg caggtaagta tcaaggttac aagacaggtt taaggagacc
     2761 aatagaaact gggcttgtcg agacagagaa gactcttgcg tttctgatag gcacctattg
     2821 gtcttactga catccacttt gcctttctct ccacaggtgt ccactcccag ttcaattaca
     2881 gctcttaagg ctagagtatt aatacgactc actatagggc tagcgatcgc catggagtgg
     2941 ccggcgcggc tctgcgggct gtgggcgctg ctgctctgcg ccggcggcgg gggcgggggc
     3001 gggggcgccg cgcctacgga aactcagcca cctgtgacaa atttgagtgt ctctgttgaa
     3061 aacctctgca cagtaatatg gacatggaat ccacccgagg gagccagctc aaattgtagt
     3121 ctatggtatt ttagtcattt tggcgacaaa caagataaga aaatagctcc ggaaactcgt
     3181 cgttcaatag aagtacccct gaatgagagg atttgtctgc aagtggggtc ccagtgtagc
     3241 accaatgaga gtgagaagcc tagcattttg gttgaaaaat gcatctcacc cccagaaggt
     3301 gatcctgagt ctgctgtgac tgagcttcaa tgcatttggc acaacctgag ctacatgaag
     3361 tgttcttggc tccctggaag gaataccagt cccgacacta actatactct ctactattgg
     3421 cacagaagcc tggaaaaaat tcatcaatgt gaaaacatct ttagagaagg ccaatacttt
     3481 ggttgttcct ttgatctgac caaagtgaag gattccagtt ttgaacaaca cagtgtccaa
     3541 ataatggtca aggataatgc aggaaaaatt aaaccatcct tcaatatagt gcctttaact
     3601 tcccgtgtga aacctgatcc tccacatatt aaaaacctct ccttccacaa tgatgaccta
     3661 tatgtgcaat gggagaatcc acagaatttt attagcagat gcctatttta tgaagtagaa
     3721 gtcaataaca gccaaactga gacacataat gttttctacg tccaagaggc taaatgtgag
     3781 aatccagaat ttgagagaaa tgtggagaat acatcttgtt tcatggtccc tggtgttctt
     3841 cctgatactt tgaacacagt cagaataaga gtcaaaacaa ataagttatg ctatgaggat
     3901 gacaaactct ggagtaattg gagccaagaa atgagtatag gtaagaagcg caattccaca
     3961 ctctacataa ccatgttact cattgttcca gtcatcgtcg caggtgcaat catagtactc
     4021 ctgctttacc taaaaaggct caagattatt atattccctc caattcctga tcctggcaag
     4081 atttttaaag aaatgtttgg agaccagaat gatgatactc tgcactggaa gaagtacgac
     4141 atctatgaga agcaaaccaa ggaggaaacc gactctgtag tgctgataga aaacctgaag
     4201 aaagcctctc aggtttctct cgagccaacc actgaggatc tgtactttca gagcgataac
     4261 gatggatccg aaatcggtac tggctttcca ttcgaccccc attatgtgga agtcctgggc
     4321 gagcgcatgc actacgtcga tgttggtccg cgcgatggca cccctgtgct gttcctgcac
     4381 ggtaacccga cctcctccta cgtgtggcgc aacatcatcc cgcatgttgc accgacccat
     4441 cgctgcattg ctccagacct gatcggtatg ggcaaatccg acaaaccaga cctgggttat
     4501 ttcttcgacg accacgtccg cttcatggat gccttcatcg aagccctggg tctggaagag
     4561 gtcgtcctgg tcattcacga ctggggctcc gctctgggtt tccactgggc caagcgcaat
     4621 ccagagcgcg tcaaaggtat tgcatttatg gagttcatcc gccctatccc gacctgggac
     4681 gaatggccag aatttgcccg cgagaccttc caggccttcc gcaccaccga cgtcggccgc
     4741 aagctgatca tcgatcagaa cgtttttatc gagggtacgc tgccgatggg tgtcgtccgc
     4801 ccgctgactg aagtcgagat ggaccattac cgcgagccgt tcctgaatcc tgttgaccgc
     4861 gagccactgt ggcgcttccc aaacgagctg ccaatcgccg gtgagccagc gaacatcgtc
     4921 gcgctggtcg aagaatacat ggactggctg caccagtccc ctgtcccgaa gctgctgttc
     4981 tggggcaccc caggcgttct gatcccaccg gccgaagccg ctcgcctggc caaaagcctg
     5041 cctaactgca aggctgtgga catcggcccg ggtctgaatc tgctgcaaga agacaacccg
     5101 gacctgatcg gcagcgagat cgcgcgctgg ctgtctactc tggagatttc cggttaatag
     5161 aattctagag tcgacctgca ggcatgcaag ctgatttggc tgctgccacc gctgagcaag
     5221 gccgcttcga gcagacatga taagatacat tgatgagttt ggacaaacca caactagaat
     5281 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     5341 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     5401 gggggagatg tgggaggttt ttttaagcaa gtaaaacctc tacaaatgtg gtaaaatctt
     5461 aattaagaat tctaatataa cttcgtatag catacattat acgaagttat ggatccactg
     5521 ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag gtgaggaact
     5581 aaacccagga ggcagatcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     5641 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     5701 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     5761 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
     5821 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     5881 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     5941 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     6001 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     6061 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     6121 aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     6181 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     6241 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     6301 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     6361 gccttctatc gccttcttga cgagttcttc tgattcgaaa tgaccgacca agcgacgccc
     6421 aaccggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     6481 caggaaagaa catgtcaaac atgagaatta caacttatat cgtatggggc tgacttcagg
     6541 tgctacattt gaagagataa attgcactga aatctagaaa tattttatct gattaataag
     6601 atgatcttc
//
