LOCUS       sequence-447060-        6865 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of secreted protein fused to 
            Fc(DAPA)-Avi-6xHis..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6865)
  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 6865)
  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-IGFBPL1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6865
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     65..84
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     90..109
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      195..783
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     684..703
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     937..954
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1052..1069
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1053..1069
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1525..1541
                     /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        1708..2087
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2088..2291
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2242..2262
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2288..2312
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2522..2540
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2522..2539
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2553..2572
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             4148..4192
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4193..4210
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4350..4367)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4356..4467
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4585..4605)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4590..4919
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4770..4905
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4832..4851
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4967..5014
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5033..5632
                     /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(5033..5052)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5529..5549
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5762..5883
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5799..5818)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5853..5872
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5936..6727,6728..6796))
                     /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     6559..6578
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(6797..6865,1..23))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atgtatttag aaaaataaac aaataggggt cagtgttaca accaattaac caattctgaa
       61 cattatcgcg agcccattta tacctgaata tggctcataa caccccttgc tcatgaccaa
      121 aatcccttaa cgtgagttac gcgcgcgtcg ttccactgag cgtcagaccc cgtagaaaag
      181 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
      241 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
      301 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
      361 ttagcccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
      421 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
      481 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
      541 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
      601 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
      661 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
      721 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
      781 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
      841 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
      901 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
      961 gaaggcgaga gtagggaact gccaggcatc aaactaagca gaaggcccct gacggatggc
     1021 ctttttgcgt ttctacaaac tctttctgtg ttgtaaaacg acggccagtc ttaagctcgg
     1081 gccccctggg cggttctgat aacgagtaat cgttaatccg caaataacgt aaaaacccgc
     1141 ttcggcgggt ttttttatgg ggggagttta gggaaagagc atttgtcaga atatttaagg
     1201 gcgcctgtca ctttgcttga tatatgagaa ttatttaacc ttataaatga gaaaaaagca
     1261 acgcacttta aataagatac gttgcttttt cgattgatga acacctataa ttaaactatt
     1321 catctattat ttatgatttt ttgtatatac aatatttcta gtttgttaaa gagaattaag
     1381 aaaataaatc tcgaaaataa taaagggaaa atcagttttt gatatcaaaa ttatacatgt
     1441 caacgataat acaaaatata atacaaacta taagatgtta tcagtattta ttatcattta
     1501 gaataaattt tgtgtcgccc ttaattgtga gcggataaca attacgagct tcatgcacag
     1561 tggcgttgac tcaatattgg ccattagcca tattattcat tggttatata gcataaatca
     1621 atattggcta ttggccattg catacgttgt atctatatca taatatgtac atttatattg
     1681 gctcatgtcc aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat
     1741 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
     1801 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
     1861 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
     1921 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
     1981 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
     2041 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     2101 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     2161 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     2221 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     2281 taagcagagc tcgtttagtg aaccgtcaga tcgcctggag acgccatcca cgctgttttg
     2341 acctccatag aagacaccgg gaccgatcca gcctccgcgg ccgggaacgg tgcattggaa
     2401 cgcggattcc ccgtgccaag agtgacgtaa gtaccgccta tagagtctat aggcccaccc
     2461 ccttggcttc gttagaacgc ggctacaatt aatacataac cttatgtatc atacacatac
     2521 gatttaggtg acactataga ataacatcca ctttgccttt ctctccacag gtgtccactc
     2581 ccaggtccaa ctgcacctcg gttctatcga aaacgcgcct gctagcgcca ccatgaagac
     2641 catcattgcc ctgagctaca tcttctgcct ggtgttcgct gcggccgcct tgggaatcag
     2701 agatgttgga ggcagacggc ccaagtgcgg accttgtaga cctgaaggat gtcccgctcc
     2761 tgctccttgt ccagctcctg gaattagcgc cctggatgag tgtggctgct gtgccagatg
     2821 tcttggagcc gaaggcgctt cttgtggtgg tagagctggt ggcagatgtg gccctggact
     2881 tgtgtgtgct tctcaagctg ctggtgccgc tcctgaaggc acaggactgt gtgtttgtgc
     2941 tcagaggggc accgtgtgtg gctccgatgg cagatcttat cctagcgtgt gcgccctgag
     3001 actgagagcc agacacacac ctagagcaca ccctggacat ctgcacaagg ccagagatgg
     3061 cccttgcgaa tttgctcctg tggtggtggt gcctcctaga agcgtgcaca atgtgacagg
     3121 cgctcaagtg ggcctgtctt gtgaagtgcg agctgtgcct acacctgtga tcacctggcg
     3181 gaaagtgaca aagagccctg agggaacaca ggccctggaa gaactgcccg gcgatcatgt
     3241 gaatatcgcc gtgcaagttc gcggcggacc ttctgatcat gaggccacag cctggattct
     3301 gatcaaccct ctgagaaaag aggacgaggg cgtctaccag tgccacgccg ctaatatggt
     3361 tggagaggcc gagagccaca gcaccgtgac agtgctggat ctgagcaagt acagaagctt
     3421 tcacttcccc gcacctgacg accggatggg cgcgccgggc ggcgacaaga cccacacctg
     3481 ccccccctgc ccagctccag aactgctggg agggccttcc gtgttcctgt tcccccccaa
     3541 gcccaaggac accctgatga tcagcaggac ccccgaggtg acctgcgtgg tggtggccgt
     3601 gtcccacgag gacccagagg tgaagttcaa ctggtacgtg gacggcgtgg aggtgcacaa
     3661 cgccaagacc aagcccagag aggagcagta caacagcacc tacagggtgg tgtccgtgct
     3721 gaccgtgctg caccaggact ggctgaacgg caaagaatac aagtgcaaag tctccaacaa
     3781 ggccctggct gccccaatcg aaaagacaat cagcaaggcc aagggccagc cacgggagcc
     3841 ccaggtgtac accctgcccc ccagccggga ggagatgacc aagaaccagg tgtccctgac
     3901 ctgtctggtg aagggcttct accccagcga tatcgccgtg gagtgggaga gcaacggcca
     3961 gcccgagaac aactacaaga ccaccccccc agtgctggac agcgacggca gcttcttcct
     4021 gtacagcaag ctgaccgtgg acaagtccag gtggcagcag ggcaacgtgt tcagctgcag
     4081 cgtgatgcac gaggccctgc acaaccacta cacccagaag tccctgagcc tgagccccgg
     4141 cggcagcggt cttaatgata ttttcgaggc tcagaagatt gagtggcatg aacaccacca
     4201 ccatcaccac taaggccggc ctgtagtcac tggcacaacg cgttacccgg taagccaatc
     4261 gggtatacac ggtcgtcata ctgcagacag ggttcttcta ctttgcaaga tagtcttgag
     4321 tagtaaaata aatagataga gaaaatcagc ctcgactgtg ccttctagtt gccagccatc
     4381 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     4441 ttcctaataa aatgaggaaa ttgcatcaca acactcaacc ctatctcggt ctattctttt
     4501 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     4561 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     4621 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     4681 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     4741 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     4801 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     4861 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     4921 agctcccggg agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc
     4981 atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa ccatgaccga
     5041 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     5101 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     5161 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     5221 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     5281 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     5341 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     5401 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     5461 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacatc
     5521 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     5581 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacacgtgct
     5641 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     5701 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     5761 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     5821 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     5881 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcattacc
     5941 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6001 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagcg
     6061 ctgcgatgat accgcgagaa ccacgctcac cggctccgga tttatcagca ataaaccagc
     6121 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     6181 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     6241 ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6301 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     6361 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     6421 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     6481 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     6541 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     6601 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     6661 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     6721 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     6781 aatgttgaat actcatattc ttcctttttc aatattattg aagcatttat cagggttatt
     6841 gtctcatgag cggatacata tttga
//
