LOCUS       sequence-447056-        7069 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 7069)
  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 7069)
  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-SIGLEC6-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7069
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     49..68
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     74..93
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      179..767
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     668..687
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     921..938
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1036..1053
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1037..1053
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1509..1525
                     /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        1692..2071
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2072..2275
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2226..2246
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2272..2296
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2506..2524
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2506..2523
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2537..2556
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             4336..4380
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4381..4398
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4538..4555)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4544..4655
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4773..4793)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4778..5107
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4958..5093
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5020..5039
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5155..5202
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5221..5820
                     /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(5221..5240)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5717..5737
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5950..6071
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5987..6006)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6041..6060
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6124..6915,6916..6984))
                     /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     6747..6766
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(6985..7069,1..7))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaacaaatag gggtcagtgt tacaaccaat taaccaattc tgaacattat cgcgagccca
       61 tttatacctg aatatggctc ataacacccc ttgctcatga ccaaaatccc ttaacgtgag
      121 ttacgcgcgc gtcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
      181 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
      241 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
      301 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttagcc caccacttca
      361 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
      421 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
      481 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
      541 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
      601 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
      661 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
      721 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
      781 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
      841 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
      901 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaaggc gagagtaggg
      961 aactgccagg catcaaacta agcagaaggc ccctgacgga tggccttttt gcgtttctac
     1021 aaactctttc tgtgttgtaa aacgacggcc agtcttaagc tcgggccccc tgggcggttc
     1081 tgataacgag taatcgttaa tccgcaaata acgtaaaaac ccgcttcggc gggttttttt
     1141 atggggggag tttagggaaa gagcatttgt cagaatattt aagggcgcct gtcactttgc
     1201 ttgatatatg agaattattt aaccttataa atgagaaaaa agcaacgcac tttaaataag
     1261 atacgttgct ttttcgattg atgaacacct ataattaaac tattcatcta ttatttatga
     1321 ttttttgtat atacaatatt tctagtttgt taaagagaat taagaaaata aatctcgaaa
     1381 ataataaagg gaaaatcagt ttttgatatc aaaattatac atgtcaacga taatacaaaa
     1441 tataatacaa actataagat gttatcagta tttattatca tttagaataa attttgtgtc
     1501 gcccttaatt gtgagcggat aacaattacg agcttcatgc acagtggcgt tgactcaata
     1561 ttggccatta gccatattat tcattggtta tatagcataa atcaatattg gctattggcc
     1621 attgcatacg ttgtatctat atcataatat gtacatttat attggctcat gtccaatatg
     1681 accgccatgt tggcattgat tattgactag ttattaatag taatcaatta cggggtcatt
     1741 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     1801 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     1861 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     1921 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     1981 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     2041 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     2101 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     2161 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     2221 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctcgttt
     2281 agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc atagaagaca
     2341 ccgggaccga tccagcctcc gcggccggga acggtgcatt ggaacgcgga ttccccgtgc
     2401 caagagtgac gtaagtaccg cctatagagt ctataggccc acccccttgg cttcgttaga
     2461 acgcggctac aattaataca taaccttatg tatcatacac atacgattta ggtgacacta
     2521 tagaataaca tccactttgc ctttctctcc acaggtgtcc actcccaggt ccaactgcac
     2581 ctcggttcta tcgaaaacgc gcctgctagc gccaccatga agaccatcat tgccctgagc
     2641 tacatcttct gcctggtgtt cgctgcggcc gcccaagaga gaagatttca gctggaaggc
     2701 cctgagagcc tgacagtgca agagggactg tgtgtgctgg tgccttgcag actgcctaca
     2761 acactgcctg ccagctacta cggctacggc tattggtttc tggaaggcgc cgatgtgccc
     2821 gtggccacca atgatcctga cgaggaagtg caagaagaga cacggggcag attccatctg
     2881 ctgtgggacc ccagaagaaa gaactgcagc ctgagcatcc gcgacgccag aagaagagat
     2941 aacgccgcct acttcttccg gctgaagtcc aagtggatga agtacggcta caccagcagc
     3001 aagctgagcg tcagagtgat ggccctgaca cacagaccca acatcagcat ccctggcaca
     3061 ctggaaagcg gacaccctag caatctgacc tgtagcgtgc catgggtttg cgagcagggc
     3121 acccctccta tctttagctg gatgagcgcc gctcctacct ctctgggacc tagaacaaca
     3181 cagagcagcg tgctgaccat cacacccaga cctcaggacc acagcaccaa cctgacctgc
     3241 caagtgacat ttcctggcgc tggcgtgacc atggaacgga caatccagct caatgtgtct
     3301 tacgcccctc agaaggtggc catcagcatt ttccagggca acagcgccgc cttcaagatc
     3361 ctgcagaata ccagcagcct gcctgtgctg gaaggacagg ctctgagact gctgtgtgac
     3421 gccgatggaa atcctcctgc tcacctgagc tggttccagg gctttcccgc tctgaatgcc
     3481 acacctatca gcaacacagg cgtgctggaa ctgcctcaag tgggatctgc cgaggaaggc
     3541 gacttcacct gtagagcaca gcaccctctg ggcagcctgc agatctctct gtctctgttc
     3601 gtgcactgga agcctgaagg cagagctggc ggagttggcg cgccgggcgg cgacaagacc
     3661 cacacctgcc ccccctgccc agctccagaa ctgctgggag ggccttccgt gttcctgttc
     3721 ccccccaagc ccaaggacac cctgatgatc agcaggaccc ccgaggtgac ctgcgtggtg
     3781 gtggccgtgt cccacgagga cccagaggtg aagttcaact ggtacgtgga cggcgtggag
     3841 gtgcacaacg ccaagaccaa gcccagagag gagcagtaca acagcaccta cagggtggtg
     3901 tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagaatacaa gtgcaaagtc
     3961 tccaacaagg ccctggctgc cccaatcgaa aagacaatca gcaaggccaa gggccagcca
     4021 cgggagcccc aggtgtacac cctgcccccc agccgggagg agatgaccaa gaaccaggtg
     4081 tccctgacct gtctggtgaa gggcttctac cccagcgata tcgccgtgga gtgggagagc
     4141 aacggccagc ccgagaacaa ctacaagacc acccccccag tgctggacag cgacggcagc
     4201 ttcttcctgt acagcaagct gaccgtggac aagtccaggt ggcagcaggg caacgtgttc
     4261 agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg
     4321 agccccggcg gcagcggtct taatgatatt ttcgaggctc agaagattga gtggcatgaa
     4381 caccaccacc atcaccacta aggccggcct gtagtcactg gcacaacgcg ttacccggta
     4441 agccaatcgg gtatacacgg tcgtcatact gcagacaggg ttcttctact ttgcaagata
     4501 gtcttgagta gtaaaataaa tagatagaga aaatcagcct cgactgtgcc ttctagttgc
     4561 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     4621 actgtccttt cctaataaaa tgaggaaatt gcatcacaac actcaaccct atctcggtct
     4681 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     4741 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     4801 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4861 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4921 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     4981 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     5041 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     5101 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
     5161 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
     5221 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     5281 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     5341 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     5401 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     5461 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     5521 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     5581 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     5641 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     5701 gagacatccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     5761 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     5821 cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     5881 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     5941 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     6001 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     6061 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     6121 tcattaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     6181 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     6241 ccccagcgct gcgatgatac cgcgagaacc acgctcaccg gctccggatt tatcagcaat
     6301 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     6361 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     6421 caacgttgtt gccatcgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     6481 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     6541 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     6601 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     6661 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6721 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     6781 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6841 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6901 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6961 gacacggaaa tgttgaatac tcatattctt cctttttcaa tattattgaa gcatttatca
     7021 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaat
//
