LOCUS       sequence-447057-        6874 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of secreted protein fused to 
            Fc(DAPA)-Avi-6xHis..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6874)
  AUTHORS   Wojtowicz WM, Vielmetter J, Fernandes RA, Siepe DH, Eastman CL, 
            Chisholm GB, Cox S, Klock H, Anderson PW, Rue SM, Miller JJ, Glaser 
            SM, Bragstad ML, Vance J, Lam AW, Lesley SA, Zinn K, Garcia KC
  TITLE     A Human IgSF Cell-Surface Interactome Reveals a Complex Network of 
            Protein-Protein Interactions.
  JOURNAL   Cell. 2020 Aug 20;182(4):1027-1043.e17. doi: 
            10.1016/j.cell.2020.07.025.
  PUBMED    32822567
REFERENCE   2  (bases 1 to 6874)
  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: 
            "10.1016/j.cell.2020.07"; journalName: "Cell"; date: "2020-08-20- 
            20"; volume: "182"; issue: "4"; pages: "1027-1043.e17"
COMMENT     Sequence Label: pD649-HAsp-IGFBP7-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6874
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     80..99
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     105..124
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      210..798
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     699..718
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     952..969
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1067..1084
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1068..1084
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1540..1556
                     /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)."
     enhancer        1723..2102
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2103..2306
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2257..2277
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2303..2327
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2537..2555
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2537..2554
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2568..2587
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             4172..4216
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4217..4234
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4374..4391)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4380..4491
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4609..4629)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4614..4943
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4794..4929
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4856..4875
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4991..5038
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5057..5656
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5057..5076)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5553..5573
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5786..5907
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5823..5842)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5877..5896
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5960..6751,6752..6820))
                     /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     6583..6602
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(6821..6874,1..38))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggtcagtg ttacaaccaa
       61 ttaaccaatt ctgaacatta tcgcgagccc atttatacct gaatatggct cataacaccc
      121 cttgctcatg accaaaatcc cttaacgtga gttacgcgcg cgtcgttcca ctgagcgtca
      181 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
      241 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
      301 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
      361 ctagtgtagc cgtagttagc ccaccacttc aagaactctg tagcaccgcc tacatacctc
      421 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
      481 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
      541 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
      601 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
      661 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
      721 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
      781 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
      841 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
      901 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
      961 gtgagcgagg aagcggaagg cgagagtagg gaactgccag gcatcaaact aagcagaagg
     1021 cccctgacgg atggcctttt tgcgtttcta caaactcttt ctgtgttgta aaacgacggc
     1081 cagtcttaag ctcgggcccc ctgggcggtt ctgataacga gtaatcgtta atccgcaaat
     1141 aacgtaaaaa cccgcttcgg cgggtttttt tatgggggga gtttagggaa agagcatttg
     1201 tcagaatatt taagggcgcc tgtcactttg cttgatatat gagaattatt taaccttata
     1261 aatgagaaaa aagcaacgca ctttaaataa gatacgttgc tttttcgatt gatgaacacc
     1321 tataattaaa ctattcatct attatttatg attttttgta tatacaatat ttctagtttg
     1381 ttaaagagaa ttaagaaaat aaatctcgaa aataataaag ggaaaatcag tttttgatat
     1441 caaaattata catgtcaacg ataatacaaa atataataca aactataaga tgttatcagt
     1501 atttattatc atttagaata aattttgtgt cgcccttaat tgtgagcgga taacaattac
     1561 gagcttcatg cacagtggcg ttgactcaat attggccatt agccatatta ttcattggtt
     1621 atatagcata aatcaatatt ggctattggc cattgcatac gttgtatcta tatcataata
     1681 tgtacattta tattggctca tgtccaatat gaccgccatg ttggcattga ttattgacta
     1741 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
     1801 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
     1861 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
     1921 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
     1981 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
     2041 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
     2101 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat
     2161 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
     2221 actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac
     2281 ggtgggaggt ctatataagc agagctcgtt tagtgaaccg tcagatcgcc tggagacgcc
     2341 atccacgctg ttttgacctc catagaagac accgggaccg atccagcctc cgcggccggg
     2401 aacggtgcat tggaacgcgg attccccgtg ccaagagtga cgtaagtacc gcctatagag
     2461 tctataggcc cacccccttg gcttcgttag aacgcggcta caattaatac ataaccttat
     2521 gtatcataca catacgattt aggtgacact atagaataac atccactttg cctttctctc
     2581 cacaggtgtc cactcccagg tccaactgca cctcggttct atcgaaaacg cgcctgctag
     2641 cgccaccatg aagaccatca ttgccctgag ctacatcttc tgcctggtgt tcgctgcggc
     2701 cgcctctagc tctgatacct gtggaccttg tgaacccgcc agctgtcctc cattgcctcc
     2761 tcttggatgt ctgctgggcg agacaagaga tgcctgcggc tgttgtccta tgtgtgccag
     2821 aggcgaaggc gaaccttgtg gcggaggcgg agctggaaga ggatattgtg cccctggcat
     2881 ggaatgcgtg aagtccagaa agcggagaaa gggcaaagct ggtgctgctg ctggtggacc
     2941 tggtgtttct ggcgtgtgtg tgtgcaagag cagataccct gtgtgtggca gcgacggcac
     3001 aacatatcca agcggctgtc agctgagagc tgcctctcag agagccgaga gtagaggcga
     3061 gaaggccatc acacaggtgt ccaagggcac atgtgaacag ggccctagca tcgtgacccc
     3121 tccaaaggac atctggaatg tgacaggcgc ccaggtgtac ctgagctgcg aagtgatcgg
     3181 aatccccaca ccagtgctga tctggaacaa agtgaagcgg ggccactacg gcgtgcagag
     3241 aacagaattg ctgcccggcg acagagacaa cctggccatt caaacaagag gcggacccga
     3301 gaagcacgaa gtgacaggat gggttctcgt gtcccctctg tctaaagagg atgccggcga
     3361 gtacgagtgc cacgcctcta attctcaagg ccaggcctct gccagcgcca agattacagt
     3421 ggtggatgcc ctgcacgaga tccccgtgaa aaaaggcgag ggcgccgaac ttggcgcgcc
     3481 gggcggcgac aagacccaca cctgcccccc ctgcccagct ccagaactgc tgggagggcc
     3541 ttccgtgttc ctgttccccc ccaagcccaa ggacaccctg atgatcagca ggacccccga
     3601 ggtgacctgc gtggtggtgg ccgtgtccca cgaggaccca gaggtgaagt tcaactggta
     3661 cgtggacggc gtggaggtgc acaacgccaa gaccaagccc agagaggagc agtacaacag
     3721 cacctacagg gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga
     3781 atacaagtgc aaagtctcca acaaggccct ggctgcccca atcgaaaaga caatcagcaa
     3841 ggccaagggc cagccacggg agccccaggt gtacaccctg ccccccagcc gggaggagat
     3901 gaccaagaac caggtgtccc tgacctgtct ggtgaagggc ttctacccca gcgatatcgc
     3961 cgtggagtgg gagagcaacg gccagcccga gaacaactac aagaccaccc ccccagtgct
     4021 ggacagcgac ggcagcttct tcctgtacag caagctgacc gtggacaagt ccaggtggca
     4081 gcagggcaac gtgttcagct gcagcgtgat gcacgaggcc ctgcacaacc actacaccca
     4141 gaagtccctg agcctgagcc ccggcggcag cggtcttaat gatattttcg aggctcagaa
     4201 gattgagtgg catgaacacc accaccatca ccactaaggc cggcctgtag tcactggcac
     4261 aacgcgttac ccggtaagcc aatcgggtat acacggtcgt catactgcag acagggttct
     4321 tctactttgc aagatagtct tgagtagtaa aataaataga tagagaaaat cagcctcgac
     4381 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     4441 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cacaacactc
     4501 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     4561 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     4621 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     4681 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     4741 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     4801 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4861 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     4921 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     4981 atcagcacgt gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa
     5041 ggtgaggaac taaaccatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga
     5101 cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca
     5161 caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac
     5221 gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt
     5281 ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat
     5341 ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc
     5401 gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca
     5461 gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg
     5521 ggtgcccgcc ttcctggaga catccgcgcc ccgcaacctc cccttctacg agcggctcgg
     5581 cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg
     5641 caagcccggt gcctgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag
     5701 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     5761 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     5821 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     5881 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     5941 cttggcgtaa tcatggtcat taccaatgct taatcagtga ggcacctatc tcagcgatct
     6001 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     6061 agggcttacc atctggcccc agcgctgcga tgataccgcg agaaccacgc tcaccggctc
     6121 cggatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     6181 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     6241 cagttaatag tttgcgcaac gttgttgcca tcgctacagg catcgtggtg tcacgctcgt
     6301 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6361 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     6421 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     6481 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     6541 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     6601 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     6661 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     6721 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     6781 aaaagggaat aagggcgaca cggaaatgtt gaatactcat attcttcctt tttcaatatt
     6841 attgaagcat ttatcagggt tattgtctca tgag
//
