LOCUS       Exported                9349 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to COMP5AP-AviTag-9xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    pD649-HAsp-FLT1-COMP5AP-AviTag-9xHis
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9349)
  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 9349)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9349
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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             4999..5022
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6682..6726
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6727..6753
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6893..6910)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6899..7010
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7128..7148)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7133..7462
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7313..7448
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7375..7394
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7510..7557
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7576..8175
                     /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(7576..7595)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     8072..8092
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    8305..8426
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8342..8361)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8396..8415
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(8479..9339)
                     /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     9102..9121
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9340..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 cggccgcctc taagctgaaa
     2761 gatcctgagc tgagcctgaa gggcacccag catattatgc aggccggaca gaccctgcat
     2821 ctgcagtgta gaggcgaagc cgctcacaag tggtccctgc ctgagatggt gtccaaagag
     2881 agcgagcggc tgagcatcac aaagagcgcc tgcggcagaa acggcaagca gttctgtagc
     2941 accctgacac tgaacacagc ccaggccaat cacaccggct tctacagctg caagtacctg
     3001 gccgtgccta ccagcaagaa gaaagaaacc gagagcgcca tctacatctt catcagcgac
     3061 accggcagac ccttcgtgga aatgtacagc gagatccccg agatcatcca catgaccgag
     3121 ggcagagagc tggtcatccc ttgcagagtg acaagcccca acatcacagt gaccctgaag
     3181 aagttccctc tggacacact gatccccgac ggcaagagaa tcatctggga cagccggaag
     3241 ggcttcatca tcagcaacgc cacctacaaa gagatcggcc tgctgacatg cgaggccaca
     3301 gtgaatggcc acctgtacaa gaccaactat ctgacccaca gacagaccaa caccatcatc
     3361 gacgtgcaga tcagcacccc tagacctgtg aagctgctga gaggacacac cctggtgctg
     3421 aattgcaccg ccaccacacc tctgaacacc agagtgcaga tgacctggtc ttaccccgac
     3481 gagaagaaca agagagccag cgtcagaagg cggatcgacc agagcaatag ccacgccaac
     3541 atcttctact ccgtgctgac catcgacaag atgcagaaca aggacaaggg cctgtacacc
     3601 tgtagagtgc ggagcggccc tagcttcaag tccgtgaata ccagcgtgca catctacgac
     3661 aaggccttca tcaccgtgaa gcaccggaag cagcaggtcc tggaaacagt ggccggcaag
     3721 aggtcctaca gactgagcat gaaggtcaaa gcctttccat ctcctgaggt cgtgtggctg
     3781 aaggatggac tgcctgccac agagaagtcc gccagatacc tgacaagagg ctacagcctg
     3841 atcatcaagg acgtgaccga agaggacgcc ggcaactaca ccatcctgct gtccatcaag
     3901 cagagcaacg tgttcaagaa cctgaccgcc actctgatcg tgaacgtgaa gccccagatc
     3961 tatgagaagg ccgtgtccag ctttcccgat cctgctctgt atcctctggg cagcagacag
     4021 atcctgacct gcacagccta tggcatccct cagcctacca tcaagtggtt ctggcacccc
     4081 tgcaaccaca accacagcga ggccagatgc gacttctgca gcaacaacga ggaaagcttc
     4141 atcctggacg ccgacagcaa catgggcaac agaatcgaga gcatcaccca gcggatggcc
     4201 attatcgagg gcaagaacaa aatggccagc acactggtgg tggccgactc cagaatcagc
     4261 ggcatctata tctgtatcgc cagcaacaaa gtgggcaccg tgggcagaaa catcagcttc
     4321 tacatcaccg atgtgcccaa cggcttccac gtgaacctgg aaaagatgcc caccgaaggc
     4381 gaggacctga agctgagctg caccgtgaac aagtttctgt accgcgacgt gacctggatt
     4441 ctgctgcgga cagtgaacaa ccggaccatg cactacagca tcagcaagca gaagatggct
     4501 atcaccaaag agcacagcat cacactgaac ctcaccatca tgaacgtgtc actgcaggac
     4561 agcggcacct acgcctgtag agccagaaat gtgtacaccg gcgaggaaat cctgcagaaa
     4621 aaagagatca ccatccggga ccaagaggcc ccttacctgc tgagaaatct gagcgatcac
     4681 accgtggcca tcagcagcag cacaacactg gattgccatg ccaatggcgt gcccgagcct
     4741 cagatcacct ggtttaagaa caaccacaag atccagcaag agcccggcat catcctcgga
     4801 cctggcagct ctaccctgtt catcgagaga gtcactgaag aggacgaggg cgtgtaccac
     4861 tgcaaggcca caaatcagaa aggcagcgtg gaaagcagcg cctacctgac agtgcagggc
     4921 accagcgaca agagcaacct ggaaggcgcg ccggaagatc aagttgatcc aagattgatt
     4981 gatggcaagg gcggatctct ggaggtgctg ttccagggac caggcggaag cggcaatccc
     5041 ggcggcgatc tggcccccca gatgctgcgc gagctgcagg agaccaatgc cgccctgcag
     5101 gacgtgcgcg agctgctgcg ccaccgcgtg aaggagatca ccttcctgaa gaacaccgtg
     5161 atggagtgcg acgcctgcgg tatgcagccc tctagaggct ccggctccat catcccagtt
     5221 gaggaggaga acccggactt ctggaaccgc gaggcagccg aggccctggg tgccgccaag
     5281 aagctgcagc ctgcacagac agccgccaag aacctcatca tcttcctggg cgatgggatg
     5341 ggggtgtcta cggtgacagc tgccaggatc ctaaaagggc agaagaagga caaactgggg
     5401 cctgagatac ccctggccat ggaccgcttc ccatatgtgg ctctgtccaa gacatacaat
     5461 gtagacaaac atgtgccaga cagtggagcc acagccacgg cctacctgtg cggggtcaag
     5521 ggcaacttcc agaccattgg cttgagtgca gccgcccgct ttaaccagtg caacacgaca
     5581 cgcggcaacg aggtcatctc cgtgatgaat cgggccaaga aagcagggaa gtcagtggga
     5641 gtggtaacca ccacacgagt gcagcacgcc tcgccagccg gcacctacgc ccacacggtg
     5701 aaccgcaact ggtactcgga cgccgacgtg cctgcctcgg cccgccagga ggggtgccag
     5761 gacatcgcta cgcagctcat ctccaacatg gacattgacg tgatcctagg tggaggccga
     5821 aagtacatgt ttcgcatggg aaccccagac cctgagtacc cagatgacta cagccaaggt
     5881 gggaccaggc tggacgggaa gaatctggtg caggaatggc tggcgaagcg ccagggtgcc
     5941 cggtatgtgt ggaaccgcac tgagctcatg caggcttccc tggacccgtc tgtgacccat
     6001 ctcatgggtc tctttgagcc tggagacatg aaatacgaga tccaccgaga ctccacactg
     6061 gacccctccc tgatggagat gacagaggct gccctgcgcc tgctgagcag gaacccccgc
     6121 ggcttcttcc tcttcgtgga gggtggtcgc atcgaccatg gtcatcatga aagcagggct
     6181 taccgggcac tgactgagac gatcatgttc gacgacgcca ttgagagggc gggccagctc
     6241 accagcgagg aggacacgct gagcctcgtc actgccgacc actcccacgt cttctccttc
     6301 ggaggctacc ccctgcgagg gagctccatc ttcgggctgg cccctggcaa ggcccgggac
     6361 aggaaggcct acacggtcct cctatacgga aacggtccag gctatgtgct caaggacggc
     6421 gcccggccgg atgttaccga gagcgagagc gggagccccg agtatcggca gcagtcagca
     6481 gtgcccctgg acgaagagac ccacgcaggc gaggacgtgg cggtgttcgc gcgcggcccg
     6541 caggcgcacc tggttcacgg cgtgcaggag cagaccttca tagcgcacgt catggccttc
     6601 gccgcctgcc tggagcccta caccgcctgc gacctggcgc cccccgccgg caccaccgac
     6661 gccgcgcacc cgggtggcgg aggactgaac gacatcttcg aggctcagaa aatcgaatgg
     6721 cacgaacatc accatcacca tcaccatcac cattaaggcc ggcctgtagt cactggcaca
     6781 acgcgttacc cggtaagcca atcgggtata cacggtcgtc atactgcaga cagggttctt
     6841 ctactttgca agatagtctt gagtagtaaa ataaatagat agagaaaatc agcctcgact
     6901 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     6961 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc acaacactca
     7021 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     7081 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     7141 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     7201 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     7261 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     7321 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     7381 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     7441 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     7501 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     7561 gtgaggaact aaaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     7621 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     7681 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     7741 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     7801 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     7861 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     7921 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     7981 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     8041 gtgcccgcct tcctggagac atccgcgccc cgcaacctcc ccttctacga gcggctcggc
     8101 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     8161 aagcccggtg cctgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
     8221 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     8281 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     8341 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     8401 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     8461 ttggcgtaat catggtcatt accaatgctt aatcagtgag gcacctatct cagcgatctg
     8521 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     8581 gggcttacca tctggcccca gcgctgcgat gataccgcga gaaccacgct caccggctcc
     8641 ggatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     8701 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     8761 agttaatagt ttgcgcaacg ttgttgccat cgctacaggc atcgtggtgt cacgctcgtc
     8821 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     8881 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     8941 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     9001 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     9061 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     9121 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     9181 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     9241 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     9301 aaagggaata agggcgacac ggaaatgttg aatactcata ttcttcctt
//
