LOCUS       sequence-304369-        8224 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 8224)
  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 8224)
  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-ISLR2_Ig1-3-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8224
                     /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             3829..3864
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             3874..3897
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5557..5601
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5602..5628
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5768..5785)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5774..5885
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6003..6023)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6008..6337
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6188..6323
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        6385..6432
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6451..7050
                     /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(6451..6470)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6947..6967
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7180..7301
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7217..7236)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7271..7290
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7354..8145,8146..8214))
                     /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     7977..7996
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8215..8224,1..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 cggccgcccc tagcgttcac
     2761 ctgtctgccg aacctccact ggaagcccct ggaacacctc tgagagccgg actggctttc
     2821 gtgctgcact gtatcgccga cggacacccc acacctagac tgcagtggca gctgcaaatc
     2881 cctggcggaa cagtggtgct ggaacctcct gttctgagcg gcgaagatga tggcgtggga
     2941 gccgaagaag gcgaaggcga aggcgacggg gatctgctga cacagacaca ggctcagacc
     3001 cctacaccag ctccagcttg gcctgctcct cctgccacac caagatttct ggccctggcc
     3061 aatggcagcc tgctggttcc tctgctgtct gccaaagagg ccggcgtcta cacatgcaga
     3121 gcccacaatg agctgggcgc caatagcacc tctatcagag tggccgtggc cgccacagga
     3181 cctccaaaac atgctcctgg tgctggcgga gagcctgatg gacaagcccc tacaagcgag
     3241 agaaagagca ccgccaaagg cagaggcaac agcgtgctgc cttctaagcc tgagggcaag
     3301 atcaaaggcc agggactcgc caaggtgtcc atcctgggag aaaccgagac tgagcccgaa
     3361 gaggatacca gcgagggcga agaggccgag gatcagattc tggctgatcc cgccgaggaa
     3421 cagagatgcg gaaacggcga tcccagcaga tacgtgtcca accacgcctt caatcagagc
     3481 gccgagctga agcctcacgt gttcgagctg ggagtgatcg ccctggatgt ggctgaaagg
     3541 gaagccagag tgcagctgac accactggct gctagatggg gacctggacc aggcggtgca
     3601 ggcggagcac ctagacctgg aagaaggcct ctgagactgc tgtatctgtg tccagctggt
     3661 ggcggagctg ctgtgcaatg gtccagagtt gaggaaggcg tgaacgccta ctggttcaga
     3721 ggactgaggc ccggcaccaa ctactctgtg tgtcttgctc tggctggcga ggcctgtcat
     3781 gtgcaagtgg tgttcagcac caagaaagag ctgccctctg gcgcgccgga agatcaagtt
     3841 gatccaagat tgattgatgg caagggcgga tctctggagg tgctgttcca gggaccaggc
     3901 ggaagcggca atcccggcgg cgatctggcc ccccagatgc tgcgcgagct gcaggagacc
     3961 aatgccgccc tgcaggacgt gcgcgagctg ctgcgccacc gcgtgaagga gatcaccttc
     4021 ctgaagaaca ccgtgatgga gtgcgacgcc tgcggtatgc agccctctag aggctccggc
     4081 tccatcatcc cagttgagga ggagaacccg gacttctgga accgcgaggc agccgaggcc
     4141 ctgggtgccg ccaagaagct gcagcctgca cagacagccg ccaagaacct catcatcttc
     4201 ctgggcgatg ggatgggggt gtctacggtg acagctgcca ggatcctaaa agggcagaag
     4261 aaggacaaac tggggcctga gatacccctg gccatggacc gcttcccata tgtggctctg
     4321 tccaagacat acaatgtaga caaacatgtg ccagacagtg gagccacagc cacggcctac
     4381 ctgtgcgggg tcaagggcaa cttccagacc attggcttga gtgcagccgc ccgctttaac
     4441 cagtgcaaca cgacacgcgg caacgaggtc atctccgtga tgaatcgggc caagaaagca
     4501 gggaagtcag tgggagtggt aaccaccaca cgagtgcagc acgcctcgcc agccggcacc
     4561 tacgcccaca cggtgaaccg caactggtac tcggacgccg acgtgcctgc ctcggcccgc
     4621 caggaggggt gccaggacat cgctacgcag ctcatctcca acatggacat tgacgtgatc
     4681 ctaggtggag gccgaaagta catgtttcgc atgggaaccc cagaccctga gtacccagat
     4741 gactacagcc aaggtgggac caggctggac gggaagaatc tggtgcagga atggctggcg
     4801 aagcgccagg gtgcccggta tgtgtggaac cgcactgagc tcatgcaggc ttccctggac
     4861 ccgtctgtga cccatctcat gggtctcttt gagcctggag acatgaaata cgagatccac
     4921 cgagactcca cactggaccc ctccctgatg gagatgacag aggctgccct gcgcctgctg
     4981 agcaggaacc cccgcggctt cttcctcttc gtggagggtg gtcgcatcga ccatggtcat
     5041 catgaaagca gggcttaccg ggcactgact gagacgatca tgttcgacga cgccattgag
     5101 agggcgggcc agctcaccag cgaggaggac acgctgagcc tcgtcactgc cgaccactcc
     5161 cacgtcttct ccttcggagg ctaccccctg cgagggagct ccatcttcgg gctggcccct
     5221 ggcaaggccc gggacaggaa ggcctacacg gtcctcctat acggaaacgg tccaggctat
     5281 gtgctcaagg acggcgcccg gccggatgtt accgagagcg agagcgggag ccccgagtat
     5341 cggcagcagt cagcagtgcc cctggacgaa gagacccacg caggcgagga cgtggcggtg
     5401 ttcgcgcgcg gcccgcaggc gcacctggtt cacggcgtgc aggagcagac cttcatagcg
     5461 cacgtcatgg ccttcgccgc ctgcctggag ccctacaccg cctgcgacct ggcgcccccc
     5521 gccggcacca ccgacgccgc gcacccgggt ggcggaggac tgaacgacat cttcgaggct
     5581 cagaaaatcg aatggcacga acatcaccat caccatcacc atcaccatta aggccggcct
     5641 gtagtcactg gcacaacgcg ttacccggta agccaatcgg gtatacacgg tcgtcatact
     5701 gcagacaggg ttcttctact ttgcaagata gtcttgagta gtaaaataaa tagatagaga
     5761 aaatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     5821 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     5881 gcatcacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     5941 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct
     6001 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     6061 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     6121 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     6181 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     6241 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta
     6301 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc
     6361 cattttcgga tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata
     6421 gtataatacg acaaggtgag gaactaaacc atgaccgagt acaagcccac ggtgcgcctc
     6481 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     6541 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     6601 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     6661 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     6721 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     6781 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     6841 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     6901 gccgagcgcg ccggggtgcc cgccttcctg gagacatccg cgccccgcaa cctccccttc
     6961 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     7021 tggtgcatga cccgcaagcc cggtgcctga cacgtgctac gagatttcga ttccaccgcc
     7081 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     7141 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     7201 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     7261 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     7321 acctctagct agagcttggc gtaatcatgg tcattaccaa tgcttaatca gtgaggcacc
     7381 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     7441 aactacgata cgggagggct taccatctgg ccccagcgct gcgatgatac cgcgagaacc
     7501 acgctcaccg gctccggatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     7561 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     7621 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccatcgcta caggcatcgt
     7681 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     7741 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     7801 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     7861 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     7921 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     7981 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     8041 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     8101 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     8161 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatattctt
     8221 cctt
//
