LOCUS       sequence-407029-        7942 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 7942)
  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 7942)
  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-LRRN2-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7942
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     124..143
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     149..168
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      254..842
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     743..762
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     996..1013
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1111..1128
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1112..1128
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1584..1600
                     /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        1767..2146
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2147..2350
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2301..2321
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2347..2371
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2581..2599
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2581..2598
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2612..2631
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             5284..5328
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5329..5346
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5486..5503)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5492..5603
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5721..5741)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5726..6055
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5906..6041
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5968..5987
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6103..6150
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6169..6768
                     /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(6169..6188)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6665..6685
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6898..7019
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6935..6954)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6989..7008
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7072..7863,7864..7932))
                     /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     7695..7714
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7933..7942,1..82))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
       61 tgtatttaga aaaataaaca aataggggtc agtgttacaa ccaattaacc aattctgaac
      121 attatcgcga gcccatttat acctgaatat ggctcataac accccttgct catgaccaaa
      181 atcccttaac gtgagttacg cgcgcgtcgt tccactgagc gtcagacccc gtagaaaaga
      241 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
      301 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
      361 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
      421 tagcccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
      481 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
      541 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
      601 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
      661 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
      721 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
      781 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
      841 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      901 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
      961 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     1021 aaggcgagag tagggaactg ccaggcatca aactaagcag aaggcccctg acggatggcc
     1081 tttttgcgtt tctacaaact ctttctgtgt tgtaaaacga cggccagtct taagctcggg
     1141 ccccctgggc ggttctgata acgagtaatc gttaatccgc aaataacgta aaaacccgct
     1201 tcggcgggtt tttttatggg gggagtttag ggaaagagca tttgtcagaa tatttaaggg
     1261 cgcctgtcac tttgcttgat atatgagaat tatttaacct tataaatgag aaaaaagcaa
     1321 cgcactttaa ataagatacg ttgctttttc gattgatgaa cacctataat taaactattc
     1381 atctattatt tatgattttt tgtatataca atatttctag tttgttaaag agaattaaga
     1441 aaataaatct cgaaaataat aaagggaaaa tcagtttttg atatcaaaat tatacatgtc
     1501 aacgataata caaaatataa tacaaactat aagatgttat cagtatttat tatcatttag
     1561 aataaatttt gtgtcgccct taattgtgag cggataacaa ttacgagctt catgcacagt
     1621 ggcgttgact caatattggc cattagccat attattcatt ggttatatag cataaatcaa
     1681 tattggctat tggccattgc atacgttgta tctatatcat aatatgtaca tttatattgg
     1741 ctcatgtcca atatgaccgc catgttggca ttgattattg actagttatt aatagtaatc
     1801 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     1861 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     1921 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     1981 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     2041 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     2101 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     2161 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     2221 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     2281 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     2341 aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     2401 cctccataga agacaccggg accgatccag cctccgcggc cgggaacggt gcattggaac
     2461 gcggattccc cgtgccaaga gtgacgtaag taccgcctat agagtctata ggcccacccc
     2521 cttggcttcg ttagaacgcg gctacaatta atacataacc ttatgtatca tacacatacg
     2581 atttaggtga cactatagaa taacatccac tttgcctttc tctccacagg tgtccactcc
     2641 caggtccaac tgcacctcgg ttctatcgaa aacgcgcctg ctagcgccac catgaagacc
     2701 atcattgccc tgagctacat cttctgcctg gtgttcgctg cggccgccgc tgttcctgtt
     2761 gttccttggc atgtgccctg tcctccacag tgcgcctgtc agatcagacc ctggtacacc
     2821 cctcggagca gctacagaga agccaccacc gtggactgca acgacctgtt tctgacagcc
     2881 gtgcctcctg ctctgcctgc cggaactcaa acactgctgc tgcagagcaa cagcatcgtg
     2941 cgcgtggacc agtctgagct gggctatctg gccaatctga ccgagctgga cctgagccag
     3001 aacagcttta gcgacgcccg ggactgcgat tttcatgccc tgcctcaact gctgagcctg
     3061 cacctggaag agaaccagct gacccggctg gaagatcact cttttgccgg actggccagc
     3121 ctgcaagagc tgtacctgaa tcacaaccag ctgtaccgga tcgcccctag agcctttagc
     3181 ggcctgagca acctgctgcg gctgcatctg aacagcaatc tgctgagagc catcgacagc
     3241 cgttggttcg agatgctgcc caacctggaa atcctgatga tcggcggcaa caaggtggac
     3301 gccatcctgg acatgaactt cagacccctg gccaacctga gaagcctggt gctggctggc
     3361 atgaacctga gagagatcag cgactacgcc ctggaaggcc tgcagagcct ggaaagcctg
     3421 tccttctacg acaatcagct ggccagagtg cctcggagag cactggaaca agtgcccggc
     3481 ctgaagttcc tggatctgaa caagaatccc ctgcagagag tcggccctgg cgacttcgct
     3541 aatatgctgc acctcaaaga gctgggcctg aacaacatgg aagaactggt gtccatcgac
     3601 aagttcgccc tggtcaatct gcccgaactg acaaagctgg acatcaccaa caatccccgg
     3661 ctgagcttca ttcaccccag agccttccac catctgcctc agatggaaac tctgatgctc
     3721 aacaacaacg ccctgagcgc cctgcaccag cagacagttg agagcctgcc taacctgcaa
     3781 gaagtgggcc tgcacggcaa ccccatcaga tgcgattgcg tgatcagatg ggccaacgcc
     3841 acaggcacca gagtgcggtt tatcgagcct cagagcaccc tgtgtgccga gcctccagat
     3901 ctgcagagac tgccagtcag agaggtgccc ttcagagaga tgaccgacca ctgcctgcct
     3961 ctgatcagcc ccagatcttt cccacctagc ctgcaggttg ccagcggcga atctatggtg
     4021 ctgcactgta gagccctggc cgagcctgag cctgagatct attgggttac accagccggc
     4081 ctgagactga caccagctca tgctggcaga cggtacagag tgtacccaga gggcacactg
     4141 gaactgagaa gagtgacagc cgaggaagcc ggactgtaca cttgcgtggc ccagaatctt
     4201 gtgggcgccg ataccaagac cgtgtccgtg gttgttggca gagcactgct ccagcctggc
     4261 agagatgaag gccagggact cgagctgaga gtgcaagaga cacaccccta ccacatcctg
     4321 ctgtcctggg tcacccctcc taataccgtg tccaccaacc tgacctggtc tagcgccagc
     4381 tctctgagag gacaaggcgc tacagctctg gctagactgc ctagaggcac ccacagctac
     4441 aacatcacca gactgctgca ggccaccgag tactgggctt gtctgcaagt ggccttcgcc
     4501 gatgctcata cccagctggc ttgtgtgtgg gccagaacaa aagaggccac cagctgtcat
     4561 agagccctcg gcgatagacc cggtggcgcg ccgggcggcg acaagaccca cacctgcccc
     4621 ccctgcccag ctccagaact gctgggaggg ccttccgtgt tcctgttccc ccccaagccc
     4681 aaggacaccc tgatgatcag caggaccccc gaggtgacct gcgtggtggt ggccgtgtcc
     4741 cacgaggacc cagaggtgaa gttcaactgg tacgtggacg gcgtggaggt gcacaacgcc
     4801 aagaccaagc ccagagagga gcagtacaac agcacctaca gggtggtgtc cgtgctgacc
     4861 gtgctgcacc aggactggct gaacggcaaa gaatacaagt gcaaagtctc caacaaggcc
     4921 ctggctgccc caatcgaaaa gacaatcagc aaggccaagg gccagccacg ggagccccag
     4981 gtgtacaccc tgccccccag ccgggaggag atgaccaaga accaggtgtc cctgacctgt
     5041 ctggtgaagg gcttctaccc cagcgatatc gccgtggagt gggagagcaa cggccagccc
     5101 gagaacaact acaagaccac ccccccagtg ctggacagcg acggcagctt cttcctgtac
     5161 agcaagctga ccgtggacaa gtccaggtgg cagcagggca acgtgttcag ctgcagcgtg
     5221 atgcacgagg ccctgcacaa ccactacacc cagaagtccc tgagcctgag ccccggcggc
     5281 agcggtctta atgatatttt cgaggctcag aagattgagt ggcatgaaca ccaccaccat
     5341 caccactaag gccggcctgt agtcactggc acaacgcgtt acccggtaag ccaatcgggt
     5401 atacacggtc gtcatactgc agacagggtt cttctacttt gcaagatagt cttgagtagt
     5461 aaaataaata gatagagaaa atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     5521 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     5581 taataaaatg aggaaattgc atcacaacac tcaaccctat ctcggtctat tcttttgatt
     5641 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     5701 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     5761 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     5821 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     5881 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     5941 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     6001 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     6061 cccgggagct tgtatatcca ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg
     6121 gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat gaccgagtac
     6181 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     6241 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     6301 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     6361 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     6421 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     6481 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     6541 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     6601 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacatccgcg
     6661 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     6721 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgaca cgtgctacga
     6781 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     6841 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     6901 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     6961 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     7021 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc attaccaatg
     7081 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     7141 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagcgctgc
     7201 gatgataccg cgagaaccac gctcaccggc tccggattta tcagcaataa accagccagc
     7261 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     7321 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     7381 catcgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     7441 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     7501 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     7561 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     7621 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     7681 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     7741 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     7801 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     7861 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     7921 ttgaatactc atattcttcc tt
//
