LOCUS       sequence-258933-        7694 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses TM287/288 in E.coli, to use as positive control in ELISA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7694)
  AUTHORS   Zimmermann I, Egloff P, Hutter CAJ, Kuhn BT, Brauer P, Newstead S, 
            Dawson RJP, Geertsma ER, Seeger MA
  TITLE     Generation of synthetic nanobodies against delicate proteins.
  JOURNAL   Nat Protoc. 2020 Apr 8. pii: 10.1038/s41596-020-0304-x. doi: 
            10.1038/s41596-020-0304-x.
  PUBMED    32269381
REFERENCE   2  (bases 1 to 7694)
  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"; journalName: "Nat Protoc.
            2020 Apr 8. pii: 10.1038/s41596-020-0304-x. doi: 
            10.1038/s41596-020-0304-x."
COMMENT     Sequence Label: pBXNH3LCA_TM287/288
FEATURES             Location/Qualifiers
     source          1..7694
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             56..85
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             86..109
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3657..3701
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3711..3740
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3756..3773
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3829..3846)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3999..4085
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4177..4204
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4223..4314
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4315..4383,4384..5175)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4533..4552)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5346..5934
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5835..5854
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6088..6105
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6120..6260
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6186..6205)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6346..6368
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6524..7402)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        join(7429..7694,1..19)
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     7636..7655
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 ctctctactg tttctccata cccgtttttt gggctaacag gaggaattaa ccatgcacca
       61 tcatcatcac catcatcatc atcatttaga agttttgttt caaggtccaa gtggttccgg
      121 cggcggtggc ggtagtaaaa cactcgccag atatttaaag ccgtactgga tatttgccgt
      181 tctcgcaccg cttttcatgg ttgtagaagt gatatgtgat ctttcacaac ccactctttt
      241 ggcacgtatt gtcgacgaag gaatcgcgcg tggagacttt tctctggttt tgaaaacagg
      301 gatcctcatg ttgatagtgg cactgatagg tgcggtggga ggaataggct gcacggtctt
      361 cgcgagctat gccagtcaga actttggagc cgacttgagg cgagatctct tcagaaaagt
      421 tctgagtttt tctatttcca atgtgaacag attccacact tcatctttga taacgcgtct
      481 caccaacgac gtcacgcagc ttcaaaactt ggtgatgatg cttctcagga tcgttgtcag
      541 agcgcctctt ctctttgtgg gtggaatagt catggcagtg tctatcaacg tgaagctttc
      601 ttccgttttg atctttttga tacctcccat cgttctcctc tttgtgtggc tcactaagaa
      661 agggaatcct ctttttcgaa agattcaaga gagcaccgac gaggtcaaca gagtcgtgag
      721 agaaaatctc ctgggtgtca gggtagtgag ggcctttcga agagaagaat acgagaatga
      781 gaactttcgg aaagcaaacg agtctctgag aagatccatc atcagtgcct tttcactcat
      841 tgttttcgca ctgccccttt tcatattcat cgtgaatatg gggatgatcg ccgtcctgtg
      901 gtttggaggg gttctcgtta gaaacaacca gatggagata ggaagcatca tggcatacac
      961 gaattacctc atgcagatca tgttctctct catgatgatc ggaaacattc tcaatttcat
     1021 tgtaagagcc agtgcttctg caaagagagt tctcgaggtg ttgaacgaaa aacctgcgat
     1081 tgaagaagcg gacaatgcct tagcacttcc aaatgtggag gggagcgttt ccttcgaaaa
     1141 tgtcgaattc agatacttcg aaaacaccga tcctgttctc agtggagtga acttctccgt
     1201 taaaccagga tcactcgttg ccgttctggg tgaaacaggg tccggaaaga gcaccctcat
     1261 gaacctcatt ccaagactca tcgatccaga acgtggacgt gtggaagtcg atgaactgga
     1321 tgttcggacg gtgaagctga aagatctgag aggtcacata tctgctgttc ctcaggaaac
     1381 cgtgctgttt tcgggaacca tcaaagagaa cctgaaatgg ggaagagaag acgccacaga
     1441 tgatgaaatc gtggaggcgg cgaagatcgc tcagatacac gatttcataa tcagtctgcc
     1501 cgaaggatac gattctcgcg tcgaaagagg ggggagaaac ttctcgggag gtcagaaaca
     1561 gagactctct atagcccgtg ctcttgtaaa aaaaccaaag gtactgatac tcgacgactg
     1621 cacgagttca gtggatccca taactgagaa aagaatactg gatgggttga agcgatacac
     1681 gaagggatgt acgaccttca tcatcactca gaagatccca acggctcttc tcgccgataa
     1741 gatcctggtc cttcatgaag ggaaagtagc gggatttggc acccacaaag agttgctcga
     1801 acactgcaaa ccctatcgag agatatatga atcccagttt ggaaacgggg tgatgaacga
     1861 tgcctgagat cagaagaagg ccacacggac caattttaga aaaacccgca ctgaaaaatc
     1921 ccacggccac gttgagaaga ctcctcggtt atctgagacc tcacactttc actttgatca
     1981 tggtctttgt tttcgtcacc gtttcttcca tacttggtgt tctctcaccg tatctcatag
     2041 gaaaaacgat cgatgttgtc ttcgttccga gaagattcga tcttcttccc agatacatgc
     2101 tgattctcgg aacgatttat gcgcttacgt ctcttctctt ctggcttcag ggaaaaatca
     2161 tgttaacgct ttcccaagat gttgtttttc gtttgagaaa ggagcttttc gagaaacttc
     2221 aaagagttcc ggtcggcttc ttcgacagaa cacctcacgg agacatcata agcagagtca
     2281 taaacgatgt ggacaatata aacaacgttc ttggaaacag tatcattcag ttcttctcgg
     2341 gaattgtgac actcgcaggt gctgtgatca tgatgttcag agtgaatgtg atcctttcac
     2401 tcgtcacact ctcaatagtt cccttgaccg ttctcatcac gcagatcgtc tccagccaaa
     2461 ccagaaaata cttctacgaa aatcaaagag tcctcggcca gctcaatggg atcatagaag
     2521 aagacatctc cggcctcact gtgataaaac tcttcacccg tgaggaaaaa gagatggaga
     2581 agttcgaccg ggtgaacgag agtctcagaa aggtgggaac gaaggctcag atcttttcag
     2641 gtgttctccc ccctctgatg aacatggtca acaacctcgg atttgccctg atcagcggtt
     2701 ttggaggatg gctcgccttg aaagacataa tcacggtggg taccatagcc actttcatag
     2761 gttactcaag gcagttcaca agaccgttga acgagctttc taaccagttc aacatgatcc
     2821 agatggcgct tgcgagtgct gaaagaatat ttgagattct cgatctggag gaagaaaaag
     2881 acgatcctga tgctgtggaa ctgagagaag tcagaggaga aatcgagttc aagaatgtct
     2941 ggttctccta cgacaagaaa aagcctgttc ttaaagacat aacctttcac ataaaaccgg
     3001 gtcagaaggt tgcactggtg ggtcccacag ggtccggaaa gacgacgatc gtgaatctgt
     3061 tgatgagatt ctacgacgtg gacagagggc agatcctcgt ggatggtata gatataagaa
     3121 aaataaagag gagttctctg agatccagca taggcatcgt ccttcaggat actattttgt
     3181 tttccaccac cgtgaaagaa aacctcaaat acggaaatcc tggcgctacc gacgaagaaa
     3241 taaaagaagc agcgaaactg acacattccg accacttcat aaagcacctg cctgagggtt
     3301 acgaaacggt cctcaccgac aacggtgaag atctgagtca gggtcaaaga cagcttcttg
     3361 ccataacaag agctttcctt gcgaatccaa agatcctgat actggacgaa gccaccagca
     3421 acgttgacac aaagacggag aaaagcatac aggcagccat gtggaaactc atggagggga
     3481 agaccagcat catcatagct cacagattga acacgataaa aaacgcagat ctgatcatcg
     3541 ttttaagaga cggagaaatt gtggagatgg gaaaacacga tgagttgatt caaaagcgag
     3601 gcttttatta cgagctcttc acaagccaat acggtctcgt tgtagaaaaa gaagcaggtc
     3661 tgaacgatat cttcgaagct cagaaaatcg aatggcacga ataataacta gaacaaaaac
     3721 tcatctcaga agaggatctg aatagcgccg tcgaccatca tcatcatcat cattgagttt
     3781 aaacggtctc cagcttggct gttttggcgg atgagagaag attttcagcc tgatacagat
     3841 taaatcagaa cgcagaagcg gtctgataaa acagaatttg cctggcggca gtagcgcggt
     3901 ggtcccacct gaccccatgc cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt
     3961 ggggtctccc catgcgagag tagggaactg ccaggcatca aataaaacga aaggctcagt
     4021 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga
     4081 caaatccgcc gggagcggat ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag
     4141 gacgcccgcc ataaactgcc aggcatcaaa ttaagcagaa ggccatcctg acggatggcc
     4201 tttttgcgtt tctacaaact cttttgttta tttttctaaa tacattcaaa tatgtatccg
     4261 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     4321 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     4381 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     4441 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     4501 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt
     4561 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     4621 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     4681 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     4741 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     4801 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     4861 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     4921 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     4981 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     5041 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     5101 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     5161 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     5221 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     5281 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     5341 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     5401 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     5461 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     5521 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     5581 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     5641 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     5701 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     5761 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     5821 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     5881 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     5941 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     6001 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     6061 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagtgccc
     6121 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat atggtgcact
     6181 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac
     6241 gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     6301 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     6361 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gcagatcaat tcgcgcgcga
     6421 aggcgaagcg gcatgcataa tgtgcctgtc aaatggacga agcagggatt ctgcaaaccc
     6481 tatgctactc cgtcaagccg tcaattgtct gattcgttac caattatgac aacttgacgg
     6541 ctacatcatt cactttttct tcacaaccgg cacggaactc gctcgggctg gccccggtgc
     6601 attttttaaa tacccgcgag aaatagagtt gatcgtcaaa accaacattg cgaccgacgg
     6661 tggcgatagg catccgggtg gtgctcaaaa gcagcttcgc ctggctgata cgttggtcct
     6721 cgcgccagct taagacgcta atccctaact gctggcggaa aagatgtgac agacgcgacg
     6781 gcgacaagca aacatgctgt gcgacgctgg cgatatcaaa attgctgtct gccaggtgat
     6841 cgctgatgta ctgacaagcc tcgcgtaccc gattatccat cggtggatgg agcgactcgt
     6901 taatcgcttc catgcgccgc agtaacaatt gctcaagcag atttatcgcc agcagctccg
     6961 aatagcgccc ttccccttgc ccggcgttaa tgatttgccc aaacaggtcg ctgaaatgcg
     7021 gctggtgcgc ttcatccggg cgaaagaacc ccgtattggc aaatattgac ggccagttaa
     7081 gccattcatg ccagtaggcg cgcggacgaa agtaaaccca ctggtgatac cattcgcgag
     7141 cctccggatg acgaccgtag tgatgaatct ctcctggcgg gaacagcaaa atatcacccg
     7201 gtcggcaaac aaattctcgt ccctgatttt tcaccacccc ctgaccgcga atggtgagat
     7261 tgagaatata acctttcatt cccagcggtc ggtcgataaa aaaatcgaga taaccgttgg
     7321 cctcaatcgg cgttaaaccc gccaccagat gggcattaaa cgagtatccc ggcagcaggg
     7381 gatcattttg cgcttcagcc atacttttca tactcccgcc attcagagaa gaaaccaatt
     7441 gtccatattg catcagacat tgccgtcact gcgtctttta ctggcacttc tcgctaacca
     7501 aaccggtaac cccgcttatt aaaagcattc tgtaacaaag cgggaccaaa gccatgacaa
     7561 aaacgcgtaa caaaagtgtc tataatcacg gcagaaaagt ccacattgat tatttgcacg
     7621 gcgtcacact ttgctatgcc atagcatttt tatccataag attagcggat cctacctgac
     7681 gctttttatc gcaa
//
