LOCUS       sequence-353482-        6815 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Baculo expression of His10 tag fused to SNAPtag and human Csk.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6815)
  AUTHORS   Hui E, Vale RD
  TITLE     In vitro membrane reconstitution of the T-cell receptor proximal 
            signaling network.
  JOURNAL   Nat Struct Mol Biol. 2014 Feb;21(2):133-42. doi: 10.1038/nsmb.2762. 
            Epub 2014 Jan 26.
  PUBMED    24463463
REFERENCE   2  (bases 1 to 6815)
  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: "10.1038/nsmb.2762";
            journalName: "Nat Struct Mol Biol"; date: "2014-02"; volume: "21"; 
            issue: "2"; pages: "133-42"
COMMENT     Sequence Label: pFastBac(His10-SNAP-Csk)
FEATURES             Location/Qualifiers
     source          1..6815
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      2..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     298..319
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        482..586
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(587..655,656..1447)
                     /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     complement(805..824)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1618..2206
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2107..2126
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2360..2377
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2511..2735
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2802..3335)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLWSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3505..3510,3511..3527,3528..3533))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        3885..3976
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     3901..3918
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4043..4072
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4094..4114
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             4127..4147
                     /codon_start=1
                     /product="tobacco vein mottling virus (TVMV) NIa protease 
                     recognition and cleavage site
                     "
                     /label=TVMV site
                     /translation="ETVRFQS"
     CDS             4184..4729
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="optimized for bacterial expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHEIKLLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYQQLAALAGNPAATAAVKTALSGNPVPILIPCHRVVSSSGAVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     polyA_signal    6305..6439
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6342..6361)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6396..6415
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  6468..6633
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt cttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
      541 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
      601 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
      661 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
      721 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
      781 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
      841 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
      901 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
      961 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     1021 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     1081 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     1141 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     1201 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1261 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     1321 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     1381 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     1441 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     1501 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     1561 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     1621 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     1681 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     1741 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     1801 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     1861 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     1921 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     1981 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     2041 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     2101 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     2161 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     2221 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     2281 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     2341 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgtg
     2401 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atagaccagc cgcgtaacct
     2461 ggcaaaatcg gttacggttg agtaataaat ggatgccctg cgtaagcggg tgtgggcgga
     2521 caataaagtc ttaaactgaa caaaatagat ctaaactatg acaataaagt cttaaactag
     2581 acagaatagt tgtaaactga aatcagtcca gttatgctgt gaaaaagcat actggacttt
     2641 tgttatggct aaagcaaact cttcattttc tgaagtgcaa attgcccgtc gtattaaaga
     2701 ggggcgtggc caagggcatg gtaaagacta tattcgcggc gttgtgacaa tttaccgaac
     2761 aactccgcgg ccgggaagcc gatctcggct tgaacgaatt gttaggtggc ggtacttggg
     2821 tcgatatcaa agtgcatcac ttcttcccgt atgcccaact ttgtatagag agccactgcg
     2881 ggatcgtcac cgtaatctgc ttgcacgtag atcacataag caccaagcgc gttggcctca
     2941 tgcttgagga gattgatgag cgcggtggca atgccctgcc tccggtgctc gccggagact
     3001 gcgagatcat agatatagat ctcactacgc ggctgctcaa acttgggcag aacgtaagcc
     3061 gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta
     3121 cggagcaagt tcccgaggta atcggagtcc ggctgatgtt gggagtaggt ggctacgtct
     3181 ccgaactcac gaccgaaaag atcaagagca gcccgcatgg atttgacttg gtcagggccg
     3241 agcctacatg tgcgaatgat gcccatactt gagccaccta actttgtttt agggcgactg
     3301 ccctgctgcg taacatcgtt gctgctccat aacatcaaac atcgacccac ggcgtaacgc
     3361 gcttgctgct tggatgcccg aggcatagac tgtacaaaaa aacagtcata acaagccatg
     3421 aaaaccgcca ctgcgccgtt accaccgctg cgttcggtca aggttctgga ccagttgcgt
     3481 gagcgcatac gctacttgca ttacagttta cgaaccgaac aggcttatgt caactgggtt
     3541 cgtgccttca tccgtttcca cggtgtgcgt cacccggcaa ccttgggcag cagcgaagtc
     3601 gaggcatttc tgtcctggct ggcgaacgag cgcaaggttt cggtctccac gcatcgtcag
     3661 gcattggcgg ccttgctgtt cttctacggc aaggtgctgt gcacggatct gccctggctt
     3721 caggagatcg gaagacctcg gccgtcgcgg cgcttgccgg tggtgctgac cccggatgaa
     3781 gtggttcgca tcctcggttt tctggaaggc gagcatcgtt tgttcgccca ggactctagc
     3841 tatagttcta gtggttggct acgtatactc cggaatatta atagatcatg gagataatta
     3901 aaatgataac catctcgcaa ataaataagt attttactgt tttcgtaaca gttttgtaat
     3961 aaaaaaacct ataaatattc cggattattc ataccgtccc accatcgggc gcggatctcg
     4021 gtccgaaacc atgtcgtact accatcacca tcaccatcac catcaccatc acgattacga
     4081 tatcccaacg accgaaaacc tgtattttca gggcgccatg ggatccgaga ctgtccgatt
     4141 ccaatctgga attcaaaggc ctacgtcgac gagctcaact agaatggaca aagattgcga
     4201 aatgaaacgt accaccctgg atagcccgct gggcaaactg gaactgagcg gctgcgaaca
     4261 gggcctgcat gaaattaaac tgctgggtaa aggcaccagc gcggccgatg cggttgaagt
     4321 tccggccccg gccgccgtgc tgggtggtcc ggaaccgctg atgcaggcga ccgcgtggct
     4381 gaacgcgtat tttcatcagc cggaagcgat tgaagaattt ccggttccgg cgctgcatca
     4441 tccggtgttt cagcaggaga gctttacccg tcaggtgctg tggaaactgc tgaaagtggt
     4501 taaatttggc gaagtgatta gctatcagca gctggcggcc ctggcgggta atccggcggc
     4561 caccgccgcc gttaaaaccg cgctgagcgg taacccggtg ccgattctga ttccgtgcca
     4621 tcgtgtggtt agctctagcg gtgcggttgg cggttatgaa ggtggtctgg cggtgaaaga
     4681 gtggctgctg gcccatgaag gtcatcgtct gggtaaaccg ggtctgggag gtagcggtag
     4741 cggtagcggt ggaggtggaa gcagcactag atcagcaata caggccgcct ggccatccgg
     4801 tacagaatgt attgccaagt acaacttcca cggcactgcc gagcaggacc tgcccttctg
     4861 caaaggagac gtgctcacca ttgtggccgt caccaaggac cccaactggt acaaagccaa
     4921 aaacaaggtg ggccgtgagg gcatcatccc agccaactac gtccagaagc gggagggcgt
     4981 gaaggcgggt accaaactca gcctcatgcc ttggttccac ggcaagatca cacgggagca
     5041 ggctgagcgg cttctgtacc cgccggagac aggcctgttc ctggtgcggg agagcaccaa
     5101 ctaccccgga gactacacgc tgtgcgtgag ctgcgacggc aaggtggagc actaccgcat
     5161 catgtaccat gccagcaagc tcagcatcga cgaggaggtg tactttgaga acctcatgca
     5221 gctggtggag cactacacct cagacgcaga tggactctgt acgcgcctca ttaaaccaaa
     5281 ggtcatggag ggcacagtgg cggcccagga tgagttctac cgcagcggct gggccctgaa
     5341 catgaaggag ctgaagctgc tgcagaccat cgggaagggg gagttcggag acgtgatgct
     5401 gggcgattac cgagggaaca aagtcgccgt caagtgcatt aagaacgacg ccactgccca
     5461 ggccttcctg gctgaagcct cagtcatgac gcaactgcgg catagcaacc tggtgcagct
     5521 cctgggcgtg atcgtggagg agaagggcgg gctctacatc gtcactgagt acatggccaa
     5581 ggggagcctt gtggactacc tgcggtctag gggtcggtca gtgctgggcg gagactgtct
     5641 cctcaagttc tcgctagatg tctgcgaggc catggaatac ctggagggca acaatttcgt
     5701 gcatcgagac ctggctgccc gcaatgtgct ggtgtctgag gacaacgtgg ccaaggtcag
     5761 cgactttggt ctcaccaagg aggcgtccag cacccaggac acgggcaagc tgccagtcaa
     5821 gtggacagcc cctgaggccc tgagagagaa gaaattctcc actaagtctg acgtgtggag
     5881 tttcggaatc cttctctggg aaatctactc ctttgggcga gtgccttatc caaggattcc
     5941 cctgaaggac gtcgtccctc gggtggagaa gggctacaag atggatgccc ccgacggctg
     6001 cccgcccgca gtctatgaag tcatgaagaa ctgctggcac ctggacgccg ccatgcggcc
     6061 ctccttccta cagctccgag agcagcttga gcacatcaaa acccacgagc tgcacctgtg
     6121 aactagtgcg gccgctttcg aatctagagc ctgcagtctc gaggcatgcg gtaccaagct
     6181 tgtcgagaag tactagagga tcataatcag ccataccaca tttgtagagg ttttacttgc
     6241 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
     6301 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     6361 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     6421 atcttatcat gtctggatct gatcactgct tgagcctagg agatccgaac cagataagtg
     6481 aaatctagtt ccaaactatt ttgtcatttt taattttcgt attagcttac gacgctacac
     6541 ccagttccca tctattttgt cactcttccc taaataatcc ttaaaaactc catttccacc
     6601 cctcccagtt cccaactatt ttgtccgccc acagcggggc atttttcttc ctgttatgtt
     6661 tttaatcaaa catcctgcca actccatgtg acaaaccgtc atcttcggct actttttctc
     6721 tgtcacagaa tgaaaatttt tctgtcatct cttcgttatt aatgtttgta attgactgaa
     6781 tatcaacgct tatttgcagc ctgaatggcg aatgg
//
