LOCUS Exported 8302 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression of cell-surface protein extracellular domain fused to Fc(DAPA)-Avi-6xHis. Protein is secreted from cells.. ACCESSION . VERSION . KEYWORDS pD649-HAsp-FLT1-Fc(DAPA)-AviTag-6xHis SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8302) 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 8302) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8302 /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 5644..5688 /codon_start=1 /product="peptide tag that allows for enzymatic biotinylation" /label=AviTag(TM) /translation="GLNDIFEAQKIEWHE" CDS 5689..5706 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(5846..5863) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 5852..5963 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(6081..6101) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 6086..6415 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 6266..6401 /label=SV40 ori /note="SV40 origin of replication" primer_bind 6328..6347 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 6463..6510 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 6529..7128 /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(6529..6548) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 7025..7045 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 7258..7379 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(7295..7314) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 7349..7368 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" CDS complement(7432..8292) /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 8055..8074 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(8293..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 ccgggcggcg acaagaccca cacctgcccc 4981 ccctgcccag ctccagaact gctgggaggg ccttccgtgt tcctgttccc ccccaagccc 5041 aaggacaccc tgatgatcag caggaccccc gaggtgacct gcgtggtggt ggccgtgtcc 5101 cacgaggacc cagaggtgaa gttcaactgg tacgtggacg gcgtggaggt gcacaacgcc 5161 aagaccaagc ccagagagga gcagtacaac agcacctaca gggtggtgtc cgtgctgacc 5221 gtgctgcacc aggactggct gaacggcaaa gaatacaagt gcaaagtctc caacaaggcc 5281 ctggctgccc caatcgaaaa gacaatcagc aaggccaagg gccagccacg ggagccccag 5341 gtgtacaccc tgccccccag ccgggaggag atgaccaaga accaggtgtc cctgacctgt 5401 ctggtgaagg gcttctaccc cagcgatatc gccgtggagt gggagagcaa cggccagccc 5461 gagaacaact acaagaccac ccccccagtg ctggacagcg acggcagctt cttcctgtac 5521 agcaagctga ccgtggacaa gtccaggtgg cagcagggca acgtgttcag ctgcagcgtg 5581 atgcacgagg ccctgcacaa ccactacacc cagaagtccc tgagcctgag ccccggcggc 5641 agcggtctta atgatatttt cgaggctcag aagattgagt ggcatgaaca ccaccaccat 5701 caccactaag gccggcctgt agtcactggc acaacgcgtt acccggtaag ccaatcgggt 5761 atacacggtc gtcatactgc agacagggtt cttctacttt gcaagatagt cttgagtagt 5821 aaaataaata gatagagaaa atcagcctcg actgtgcctt ctagttgcca gccatctgtt 5881 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc 5941 taataaaatg aggaaattgc atcacaacac tcaaccctat ctcggtctat tcttttgatt 6001 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat 6061 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc 6121 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa 6181 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac 6241 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc 6301 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc 6361 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct 6421 cccgggagct tgtatatcca ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg 6481 gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat gaccgagtac 6541 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc 6601 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag 6661 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg 6721 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg 6781 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc 6841 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc 6901 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg 6961 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacatccgcg 7021 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg 7081 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgaca cgtgctacga 7141 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac 7201 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac 7261 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat 7321 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat 7381 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc attaccaatg 7441 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg 7501 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagcgctgc 7561 gatgataccg cgagaaccac gctcaccggc tccggattta tcagcaataa accagccagc 7621 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa 7681 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc 7741 catcgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg 7801 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc 7861 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat 7921 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg 7981 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc 8041 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg 8101 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat 8161 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg 8221 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg 8281 ttgaatactc atattcttcc tt //