LOCUS       sequence-302905-        7654 bp    DNA     circular SYN 15-APR-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 7654)
  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 7654)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-BCAM-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7654
                     /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             4996..5040
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5041..5058
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5198..5215)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5204..5315
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5433..5453)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5438..5767
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5618..5753
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5680..5699
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5815..5862
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5881..6480
                     /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(5881..5900)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6377..6397
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6610..6731
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6647..6666)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6701..6720
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6784..7575,7576..7644))
                     /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     7407..7426
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7645..7654,1..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 cggccgccga agttcgactg
     2761 tctgttcctc cactggtgga agtgatgcgg ggcaagtctg tgatcctgga ttgcacacct
     2821 accggcacac acgaccacta catgctggaa tggtttctga ccgacagaag cggcgctaga
     2881 cctagactgg cctctgccga aatgcagggc tctgagctgc aagtgaccat gcacgatacc
     2941 agaggcagaa gccctcctta ccagctggat tctcagggca gactggtgct ggctgaagcc
     3001 caagttggcg acgagaggga ttacgtgtgt gtcgttagag ccggcgctgc cggaacagct
     3061 gaagccacag ctagactgaa cgtgttcgcc aagcctgagg ccacagaggt gtccccaaac
     3121 aagggcacac tgagcgtgat ggaagatagc gcccaagaga tcgccacctg taacagcaga
     3181 aacggcaacc ccgctcctaa gatcacctgg tacagaaatg gccagcggct ggaagtgccc
     3241 gtggaaatga atcctgaggg ctacatgacc agcagaaccg tgcgtgaagc ctctggcctg
     3301 ctgagcctga caagcacact gtacctgcgg ctgagaaagg acgacagaga tgccagcttt
     3361 cactgcgccg ctcactactc tctgcctgaa ggcagacatg gcaggctgga cagccctacc
     3421 tttcacctga cactgcacta ccctaccgag cacgtgcagt tttgggtcgg atctccatct
     3481 acacccgccg gatgggtccg agaaggcgat acagttcaac tgctgtgtag aggcgacggc
     3541 agcccatctc ctgagtacac cctgttcaga ctgcaagacg agcaagagga agtgctgaac
     3601 gtcaacctgg aaggcaacct gactctggaa ggcgtgacaa gaggccagag cggcacatat
     3661 ggctgcagag tggaagatta cgacgccgcc gatgacgtgc agctgagcaa gacactggaa
     3721 ctgagagtgg cctatctgga ccctctggaa ctgtccgagg gcaaagttct gagcctgcct
     3781 ctgaatagca gcgccgtcgt gaattgctct gtgcacggat tgcccacacc agctctgaga
     3841 tggaccaagg atagcacccc tctcggagat ggccctatgc tgtctctgag cagcatcacc
     3901 ttcgacagca acggcacata cgtgtgcgag gcttctctgc ctacagtgcc tgtgctgagc
     3961 agaacccaga acttcacact gctggtgcag ggcagccctg agctgaaaac agccgagatt
     4021 gagcctaagg ccgacggctc ttggagagag ggcgacgaag tcacactgat ctgtagcgcc
     4081 agaggacacc ccgatccaaa gctgtcttgg agccagcttg gaggaagccc tgccgaacct
     4141 attcctggca gacaaggatg ggtgtccagc tctctgaccc tgaaagtgac aagcgccctg
     4201 agcagagatg gcatcagctg cgaagccagc aatccccacg gaaacaagcg gcacgtgttc
     4261 cactttggca ccgtgtctcc acagaccagc caagctggcg cgccgggcgg cgacaagacc
     4321 cacacctgcc ccccctgccc agctccagaa ctgctgggag ggccttccgt gttcctgttc
     4381 ccccccaagc ccaaggacac cctgatgatc agcaggaccc ccgaggtgac ctgcgtggtg
     4441 gtggccgtgt cccacgagga cccagaggtg aagttcaact ggtacgtgga cggcgtggag
     4501 gtgcacaacg ccaagaccaa gcccagagag gagcagtaca acagcaccta cagggtggtg
     4561 tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagaatacaa gtgcaaagtc
     4621 tccaacaagg ccctggctgc cccaatcgaa aagacaatca gcaaggccaa gggccagcca
     4681 cgggagcccc aggtgtacac cctgcccccc agccgggagg agatgaccaa gaaccaggtg
     4741 tccctgacct gtctggtgaa gggcttctac cccagcgata tcgccgtgga gtgggagagc
     4801 aacggccagc ccgagaacaa ctacaagacc acccccccag tgctggacag cgacggcagc
     4861 ttcttcctgt acagcaagct gaccgtggac aagtccaggt ggcagcaggg caacgtgttc
     4921 agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc cctgagcctg
     4981 agccccggcg gcagcggtct taatgatatt ttcgaggctc agaagattga gtggcatgaa
     5041 caccaccacc atcaccacta aggccggcct gtagtcactg gcacaacgcg ttacccggta
     5101 agccaatcgg gtatacacgg tcgtcatact gcagacaggg ttcttctact ttgcaagata
     5161 gtcttgagta gtaaaataaa tagatagaga aaatcagcct cgactgtgcc ttctagttgc
     5221 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     5281 actgtccttt cctaataaaa tgaggaaatt gcatcacaac actcaaccct atctcggtct
     5341 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     5401 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     5461 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     5521 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     5581 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     5641 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     5701 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     5761 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
     5821 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
     5881 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     5941 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     6001 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     6061 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     6121 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     6181 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     6241 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     6301 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     6361 gagacatccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     6421 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     6481 cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     6541 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     6601 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     6661 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     6721 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     6781 tcattaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     6841 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     6901 ccccagcgct gcgatgatac cgcgagaacc acgctcaccg gctccggatt tatcagcaat
     6961 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     7021 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     7081 caacgttgtt gccatcgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     7141 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     7201 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     7261 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     7321 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     7381 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     7441 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     7501 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     7561 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     7621 gacacggaaa tgttgaatac tcatattctt cctt
//
