LOCUS       sequence-303109-        8734 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein partial 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 8734)
  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 8734)
  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-SDK2(Trunc2)-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8734
                     /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             6076..6120
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6121..6138
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6278..6295)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6284..6395
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6513..6533)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6518..6847
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6698..6833
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6760..6779
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6895..6942
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6961..7560
                     /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(6961..6980)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7457..7477
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7690..7811
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7727..7746)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7781..7800
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7864..8655,8656..8724))
                     /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     8487..8506
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8725..8734,1..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 cggccgccca agatgatgtg
     2761 tccccttact ttaagaccga gcctgtgcgg acccaggtcc accttgaggg aaatagactg
     2821 gtgctgacct gcatggccga aggctcttgg cctctggaat tcaagtggct gcacaacaac
     2881 agagagctga ccaagttcag cctggaatac cggtacatga tcaccagcct ggatagaacc
     2941 cacgccggct tctacagatg catcgtgcgg aatagaatgg gcgccctgct gcagagacag
     3001 acagaggtgc aagtggccta catgggcagc ttcgaggaag gcgagaaaca ccagtctgtg
     3061 tctcatggcg aggccgccgt gatcagagca cctagaattg ccagctttcc ccagcctcaa
     3121 gtgacctggt tcagagatgg ccggaagatc cctcctagca gcagaatcgc catcacactg
     3181 gaaaacaccc tggtcatcct gagcacagtg gcccctgatg ccggcagata ttacgtgcag
     3241 gccgtgaacg acaagaacgg cgacaacaag accagccagc ctatcacact gaccgtggaa
     3301 aatgttggcg gccctgccga tcctatcgct cccacaatca tcatccctcc aaagaacaca
     3361 agcgtggtgg ccggcacaag cgaagtgaca ctggaatgcg tggccaacgc cagacctctg
     3421 atcaagctgc acatcatttg gaagaaagac ggcgtgctgc tgagcggcgg catcagcgat
     3481 cataacagac ggctgacaat ccccaatcct acaggctccg atgccggcta ctatgagtgt
     3541 gaagccgtgc tgagaagcag cagcgtgcca tctgttgtca gaggcgccta cctgtctgtg
     3601 ctggaacctc ctcagttcgt gaaagagccc gagagacaca tcaccgccga gatggaaaag
     3661 gtggtggata tcccttgtca ggccaagggc gtgccaccac ctagcatcac atggtacaag
     3721 gacgccgccg tggtggaagt ggaaaagctg accagattcc ggcagagaaa cgatggcggc
     3781 ctgcagattt ctggcctggt gcctgatgat accggcatgt tccagtgctt cgccagaaat
     3841 gcagccggcg aggtgcagac aagcacatac ctggccgtga cctctatcgc ccctaacatc
     3901 acaagaggcc ctctggacag caccgtgatc gatggaatgt ctgtggtgct ggcctgtgaa
     3961 acctctggcg cccctagacc tgccattacc tggcagaagg gcgagagaat tctggccagt
     4021 ggaagcgtgc agctgcctag attcacacct ctggaaagcg gcagcctgct gatctcccct
     4081 acacacatct ctgacgccgg cacctacacc tgtctggcca caaatagtag aggcgtggac
     4141 gaagccagcg ccgatctggt tgtttgggcc agaaccagaa tcaccaagcc tccacaggac
     4201 cagagcgtga tcaagggcac acaggcctct atggtttgcg gcgtgacaca cgaccctaga
     4261 gtgaccatcc ggtacatctg ggagaaagat ggggccacac tgggcaccga gtctcaccct
     4321 agaatcagac tggacagaaa cggctccctg cacatctctc agacttggag cggcgacatc
     4381 ggcacataca catgcagagt gatcagcgct ggcggcaacg acagtagatc tgcccacctg
     4441 agagtcagac agctgcctca cgctcctgaa caccctgtgg ctaccctgtc cacagtggaa
     4501 agaagggcca tcaacctgac ctggaccaag cctttcgacg gcaatagccc tctgatccgc
     4561 tacatcctgg aaatgagcga gaacaacgcc ccttggacag tgctgctggc ctctgtggac
     4621 cctaaggcca cctctgtgac agtgaaagga ctggtgccag ccagaagcta ccagttcaga
     4681 ctgtgcgccg tgaatgatgt cggcaagggc cagttcagca aggacaccga aagagtgtcc
     4741 ctgcctgagg aacctccaac cgctcctcct cagaatgtga tcgccagcgg caggaccaac
     4801 cagagcatca tgattcagtg gcagccacct cctgagagcc accagaatgg catcctgaag
     4861 ggctacatca tcagatactg cctggccgga ctgcccgtgg gctatcagtt caagaacatc
     4921 accgacgccg acgtgaacaa cctgctgctg gaagatctga tcatctggac caactacgag
     4981 atcgaggtgg ccgcctacaa ttctgccggc ctgggcgtgt acagcagcaa agtgacagaa
     5041 tggacactgc agggcgtccc aacagtgcct cctggcaatg tgcatgccga ggccaccaat
     5101 agcaccacca tcagattcac ctggaacgcc ccatctccac agttcatcaa cggcatcaac
     5161 cagggctaca agctgatcgc ctgggagcct gagcaagagg aagaggtcac aatggtcacc
     5221 gccaggccta acttccagga tagcatccac gtgggcttcg tgtccggcct gaagaagttc
     5281 accgagtact tcaccagcgt gctgtgcttc acaacacctg gcgacggccc tagatctaca
     5341 cctcagctcg tgcggacaca cgaggatgtt cccggtggcg cgccgggcgg cgacaagacc
     5401 cacacctgcc ccccctgccc agctccagaa ctgctgggag ggccttccgt gttcctgttc
     5461 ccccccaagc ccaaggacac cctgatgatc agcaggaccc ccgaggtgac ctgcgtggtg
     5521 gtggccgtgt cccacgagga cccagaggtg aagttcaact ggtacgtgga cggcgtggag
     5581 gtgcacaacg ccaagaccaa gcccagagag gagcagtaca acagcaccta cagggtggtg
     5641 tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagaatacaa gtgcaaagtc
     5701 tccaacaagg ccctggctgc cccaatcgaa aagacaatca gcaaggccaa gggccagcca
     5761 cgggagcccc aggtgtacac cctgcccccc agccgggagg agatgaccaa gaaccaggtg
     5821 tccctgacct gtctggtgaa gggcttctac cccagcgata tcgccgtgga gtgggagagc
     5881 aacggccagc ccgagaacaa ctacaagacc acccccccag tgctggacag cgacggcagc
     5941 ttcttcctgt acagcaagct gaccgtggac aagtccaggt ggcagcaggg caacgtgttc
     6001 agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg
     6061 agccccggcg gcagcggtct taatgatatt ttcgaggctc agaagattga gtggcatgaa
     6121 caccaccacc atcaccacta aggccggcct gtagtcactg gcacaacgcg ttacccggta
     6181 agccaatcgg gtatacacgg tcgtcatact gcagacaggg ttcttctact ttgcaagata
     6241 gtcttgagta gtaaaataaa tagatagaga aaatcagcct cgactgtgcc ttctagttgc
     6301 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     6361 actgtccttt cctaataaaa tgaggaaatt gcatcacaac actcaaccct atctcggtct
     6421 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     6481 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     6541 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     6601 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     6661 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     6721 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     6781 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     6841 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
     6901 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
     6961 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     7021 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     7081 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     7141 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     7201 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     7261 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     7321 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     7381 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     7441 gagacatccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     7501 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     7561 cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     7621 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     7681 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     7741 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     7801 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     7861 tcattaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     7921 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     7981 ccccagcgct gcgatgatac cgcgagaacc acgctcaccg gctccggatt tatcagcaat
     8041 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     8101 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     8161 caacgttgtt gccatcgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     8221 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     8281 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     8341 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     8401 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     8461 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     8521 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     8581 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     8641 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     8701 gacacggaaa tgttgaatac tcatattctt cctt
//
