LOCUS       sequence-433178-        6772 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to Fc(DAPA)-Avi-6xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6772)
  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 6772)
  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-KIR2DP1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6772
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     107..126
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     132..151
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      237..825
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     726..745
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     979..996
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1094..1111
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1095..1111
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1567..1583
                     /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        1750..2129
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2130..2333
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2284..2304
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2330..2354
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2564..2582
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2564..2581
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2595..2614
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             4097..4141
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4142..4159
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4299..4316)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4305..4416
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4534..4554)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4539..4868
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4719..4854
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4781..4800
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4916..4963
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4982..5581
                     /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(4982..5001)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5478..5498
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5711..5832
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5748..5767)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5802..5821
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5885..6676,6677..6745))
                     /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     6508..6527
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(6746..6772,1..65))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
       61 acaaataggg gtcagtgtta caaccaatta accaattctg aacattatcg cgagcccatt
      121 tatacctgaa tatggctcat aacacccctt gctcatgacc aaaatccctt aacgtgagtt
      181 acgcgcgcgt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
      241 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
      301 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
      361 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttagccca ccacttcaag
      421 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
      481 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
      541 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
      601 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
      661 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
      721 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
      781 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
      841 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
      901 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
      961 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaaggcga gagtagggaa
     1021 ctgccaggca tcaaactaag cagaaggccc ctgacggatg gcctttttgc gtttctacaa
     1081 actctttctg tgttgtaaaa cgacggccag tcttaagctc gggccccctg ggcggttctg
     1141 ataacgagta atcgttaatc cgcaaataac gtaaaaaccc gcttcggcgg gtttttttat
     1201 ggggggagtt tagggaaaga gcatttgtca gaatatttaa gggcgcctgt cactttgctt
     1261 gatatatgag aattatttaa ccttataaat gagaaaaaag caacgcactt taaataagat
     1321 acgttgcttt ttcgattgat gaacacctat aattaaacta ttcatctatt atttatgatt
     1381 ttttgtatat acaatatttc tagtttgtta aagagaatta agaaaataaa tctcgaaaat
     1441 aataaaggga aaatcagttt ttgatatcaa aattatacat gtcaacgata atacaaaata
     1501 taatacaaac tataagatgt tatcagtatt tattatcatt tagaataaat tttgtgtcgc
     1561 ccttaattgt gagcggataa caattacgag cttcatgcac agtggcgttg actcaatatt
     1621 ggccattagc catattattc attggttata tagcataaat caatattggc tattggccat
     1681 tgcatacgtt gtatctatat cataatatgt acatttatat tggctcatgt ccaatatgac
     1741 cgccatgttg gcattgatta ttgactagtt attaatagta atcaattacg gggtcattag
     1801 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
     1861 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
     1921 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
     1981 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     2041 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     2101 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     2161 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     2221 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
     2281 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag
     2341 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc
     2401 gggaccgatc cagcctccgc ggccgggaac ggtgcattgg aacgcggatt ccccgtgcca
     2461 agagtgacgt aagtaccgcc tatagagtct ataggcccac ccccttggct tcgttagaac
     2521 gcggctacaa ttaatacata accttatgta tcatacacat acgatttagg tgacactata
     2581 gaataacatc cactttgcct ttctctccac aggtgtccac tcccaggtcc aactgcacct
     2641 cggttctatc gaaaacgcgc ctgctagcgc caccatgaag accatcattg ccctgagcta
     2701 catcttctgc ctggtgttcg ctgcggccgc ccatgaaggc gtgcacagaa aaccttctct
     2761 gctggcccat cctggacctc tggtcaagag cgaagagaca gtgatcctgc agtgttggag
     2821 cgacgtgcgg ttccagcact ttctgctgca cagagagggc aagttcaagg acacactgca
     2881 cctgatcgga gagcaccacg atggcgtgtc caaggccaat ttcagcatcg gccccatgat
     2941 gcaggatctg gccggcacat acagatgcta cggctctgtg acacacagcc cctatcagct
     3001 gtctgcccct agcgatcccc tggatatcgt gatcaccggc ctgtacgaga agcctagcct
     3061 gtctgctcag cctggaccta cagttctggc tggcgaaagc gtgaccctga gctgtagcag
     3121 cagatccagc tacgacatgt accacctgtc cagagaaggc gaggcccacg agagaagatt
     3181 cagcgccgga cctaaagtga acggcacctt ccaggccgat tttccactgg gacctgccac
     3241 acacggcggc acctatagat gctttggcag cttcagagac agcccttacg agtggtccaa
     3301 cagcagcgat cctctgctgg tgtctgtgac cggcaatccc agcaatagct ggccttctcc
     3361 aaccgagcct agcagcaaga caggcaaccc cagacatggc gcgccgggcg gcgacaagac
     3421 ccacacctgc cccccctgcc cagctccaga actgctggga gggccttccg tgttcctgtt
     3481 cccccccaag cccaaggaca ccctgatgat cagcaggacc cccgaggtga cctgcgtggt
     3541 ggtggccgtg tcccacgagg acccagaggt gaagttcaac tggtacgtgg acggcgtgga
     3601 ggtgcacaac gccaagacca agcccagaga ggagcagtac aacagcacct acagggtggt
     3661 gtccgtgctg accgtgctgc accaggactg gctgaacggc aaagaataca agtgcaaagt
     3721 ctccaacaag gccctggctg ccccaatcga aaagacaatc agcaaggcca agggccagcc
     3781 acgggagccc caggtgtaca ccctgccccc cagccgggag gagatgacca agaaccaggt
     3841 gtccctgacc tgtctggtga agggcttcta ccccagcgat atcgccgtgg agtgggagag
     3901 caacggccag cccgagaaca actacaagac caccccccca gtgctggaca gcgacggcag
     3961 cttcttcctg tacagcaagc tgaccgtgga caagtccagg tggcagcagg gcaacgtgtt
     4021 cagctgcagc gtgatgcacg aggccctgca caaccactac acccagaagt ccctgagcct
     4081 gagccccggc ggcagcggtc ttaatgatat tttcgaggct cagaagattg agtggcatga
     4141 acaccaccac catcaccact aaggccggcc tgtagtcact ggcacaacgc gttacccggt
     4201 aagccaatcg ggtatacacg gtcgtcatac tgcagacagg gttcttctac tttgcaagat
     4261 agtcttgagt agtaaaataa atagatagag aaaatcagcc tcgactgtgc cttctagttg
     4321 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     4381 cactgtcctt tcctaataaa atgaggaaat tgcatcacaa cactcaaccc tatctcggtc
     4441 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     4501 atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa
     4561 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4621 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     4681 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     4741 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     4801 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     4861 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga
     4921 caattaatca tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac
     4981 catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt
     5041 acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga
     5101 ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga
     5161 catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga
     5221 gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg
     5281 ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga
     5341 gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg
     5401 cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct
     5461 ggagacatcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc
     5521 cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg
     5581 acacgtgcta cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     5641 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     5701 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5761 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5821 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     5881 gtcattacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5941 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     6001 gccccagcgc tgcgatgata ccgcgagaac cacgctcacc ggctccggat ttatcagcaa
     6061 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     6121 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     6181 gcaacgttgt tgccatcgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     6241 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     6301 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     6361 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     6421 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     6481 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     6541 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6601 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6661 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6721 cgacacggaa atgttgaata ctcatattct tcctttttca atattattga ag
//
