LOCUS       sequence-303693-        8014 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to COMP5AP-AviTag-9xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8014)
  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 8014)
  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-NEGR1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8014
                     /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             3619..3654
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             3664..3687
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5347..5391
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5392..5418
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5558..5575)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5564..5675
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5793..5813)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5798..6127
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5978..6113
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6040..6059
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6175..6222
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6241..6840
                     /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(6241..6260)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6737..6757
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6970..7091
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7007..7026)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7061..7080
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7144..7935,7936..8004))
                     /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     7767..7786
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8005..8014,1..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 cggccgccgt tgattttcct
     2761 tgggccgctg tggacaacat gatggtccga aaaggcgaca ccgccgtgct gagatgttat
     2821 ctggaagatg gcgccagcaa aggcgcctgg ctgaacagaa gctctatcat ctttgccggc
     2881 ggagacaagt ggtccgtcga tcctagagtg tccatcagca ccctgaacaa gcgggactac
     2941 agcctgcaga tccagaacgt ggacgtgacc gacgatggcc cttacacctg ttctgtgcag
     3001 acccagcaca cccctagaac catgcaggtc cacctgacag tccaggtgcc acctaagatc
     3061 tacgacatca gcaacgacat gaccgtgaac gagggcacca atgtgaccct gacatgtctg
     3121 gccacaggca agcctgagcc tagcatctct tggagacaca tcagccccag cgccaagcct
     3181 ttcgagaatg gccagtacct ggacatctac ggcatcacca gagatcaggc cggcgagtat
     3241 gagtgctctg ccgagaacga tgtgtcattc cccgacgtgc ggaaagtgaa ggtggtggtc
     3301 aatttcgccc ctaccattca agagatcaag agcggcaccg tgacacctgg cagatctggc
     3361 ctgattagat gtgaaggtgc tggcgtgcca cctcctgcct ttgagtggta taagggcgag
     3421 aagaagctgt tcaacggcca gcagggcatc atcatccaga acttcagcac ccggtccatc
     3481 ctgaccgtga ccaacgtgac acaagagcac ttcggcaact acacctgtgt ggccgccaac
     3541 aagctgggca ccacaaatgc ttccctgcct ctgaatcctc ctagcacagc ccagtatggc
     3601 atcaccggtg gcgcgccgga agatcaagtt gatccaagat tgattgatgg caagggcgga
     3661 tctctggagg tgctgttcca gggaccaggc ggaagcggca atcccggcgg cgatctggcc
     3721 ccccagatgc tgcgcgagct gcaggagacc aatgccgccc tgcaggacgt gcgcgagctg
     3781 ctgcgccacc gcgtgaagga gatcaccttc ctgaagaaca ccgtgatgga gtgcgacgcc
     3841 tgcggtatgc agccctctag aggctccggc tccatcatcc cagttgagga ggagaacccg
     3901 gacttctgga accgcgaggc agccgaggcc ctgggtgccg ccaagaagct gcagcctgca
     3961 cagacagccg ccaagaacct catcatcttc ctgggcgatg ggatgggggt gtctacggtg
     4021 acagctgcca ggatcctaaa agggcagaag aaggacaaac tggggcctga gatacccctg
     4081 gccatggacc gcttcccata tgtggctctg tccaagacat acaatgtaga caaacatgtg
     4141 ccagacagtg gagccacagc cacggcctac ctgtgcgggg tcaagggcaa cttccagacc
     4201 attggcttga gtgcagccgc ccgctttaac cagtgcaaca cgacacgcgg caacgaggtc
     4261 atctccgtga tgaatcgggc caagaaagca gggaagtcag tgggagtggt aaccaccaca
     4321 cgagtgcagc acgcctcgcc agccggcacc tacgcccaca cggtgaaccg caactggtac
     4381 tcggacgccg acgtgcctgc ctcggcccgc caggaggggt gccaggacat cgctacgcag
     4441 ctcatctcca acatggacat tgacgtgatc ctaggtggag gccgaaagta catgtttcgc
     4501 atgggaaccc cagaccctga gtacccagat gactacagcc aaggtgggac caggctggac
     4561 gggaagaatc tggtgcagga atggctggcg aagcgccagg gtgcccggta tgtgtggaac
     4621 cgcactgagc tcatgcaggc ttccctggac ccgtctgtga cccatctcat gggtctcttt
     4681 gagcctggag acatgaaata cgagatccac cgagactcca cactggaccc ctccctgatg
     4741 gagatgacag aggctgccct gcgcctgctg agcaggaacc cccgcggctt cttcctcttc
     4801 gtggagggtg gtcgcatcga ccatggtcat catgaaagca gggcttaccg ggcactgact
     4861 gagacgatca tgttcgacga cgccattgag agggcgggcc agctcaccag cgaggaggac
     4921 acgctgagcc tcgtcactgc cgaccactcc cacgtcttct ccttcggagg ctaccccctg
     4981 cgagggagct ccatcttcgg gctggcccct ggcaaggccc gggacaggaa ggcctacacg
     5041 gtcctcctat acggaaacgg tccaggctat gtgctcaagg acggcgcccg gccggatgtt
     5101 accgagagcg agagcgggag ccccgagtat cggcagcagt cagcagtgcc cctggacgaa
     5161 gagacccacg caggcgagga cgtggcggtg ttcgcgcgcg gcccgcaggc gcacctggtt
     5221 cacggcgtgc aggagcagac cttcatagcg cacgtcatgg ccttcgccgc ctgcctggag
     5281 ccctacaccg cctgcgacct ggcgcccccc gccggcacca ccgacgccgc gcacccgggt
     5341 ggcggaggac tgaacgacat cttcgaggct cagaaaatcg aatggcacga acatcaccat
     5401 caccatcacc atcaccatta aggccggcct gtagtcactg gcacaacgcg ttacccggta
     5461 agccaatcgg gtatacacgg tcgtcatact gcagacaggg ttcttctact ttgcaagata
     5521 gtcttgagta gtaaaataaa tagatagaga aaatcagcct cgactgtgcc ttctagttgc
     5581 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     5641 actgtccttt cctaataaaa tgaggaaatt gcatcacaac actcaaccct atctcggtct
     5701 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     5761 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     5821 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     5881 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     5941 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     6001 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     6061 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     6121 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
     6181 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
     6241 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     6301 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     6361 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     6421 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     6481 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     6541 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     6601 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     6661 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     6721 gagacatccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     6781 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     6841 cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     6901 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     6961 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     7021 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     7081 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     7141 tcattaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     7201 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     7261 ccccagcgct gcgatgatac cgcgagaacc acgctcaccg gctccggatt tatcagcaat
     7321 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     7381 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     7441 caacgttgtt gccatcgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     7501 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     7561 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     7621 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     7681 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     7741 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     7801 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     7861 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     7921 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     7981 gacacggaaa tgttgaatac tcatattctt cctt
//
