LOCUS Exported 7147 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-IL1RAP-Fc(DAPA)-AviTag-6xHis SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7147) 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 7147) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7147 /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 4489..4533 /codon_start=1 /product="peptide tag that allows for enzymatic biotinylation" /label=AviTag(TM) /translation="GLNDIFEAQKIEWHE" CDS 4534..4551 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(4691..4708) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 4697..4808 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(4926..4946) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 4931..5260 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 5111..5246 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5173..5192 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 5308..5355 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 5374..5973 /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(5374..5393) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 5870..5890 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 6103..6224 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(6140..6159) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 6194..6213 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" CDS complement(6277..7137) /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 6900..6919 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7138..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 tgagagatgt 2761 gatgattggg gcctcgacac catgcggcag atccaggtgt tcgaggatga gcccgccaga 2821 atcaagtgcc ctctgttcga gcacttcctg aagttcaact acagcacagc ccacagcgcc 2881 ggactgaccc tgatctggta ttggacccgg caggaccgcg acctggaaga acccatcaac 2941 ttcagactgc ccgagaaccg gatcagcaaa gaaaaggacg tgctgtggtt cagacccaca 3001 ctgctgaacg acaccggcaa ctacacctgt atgctgcgga acaccaccta ctgcagcaag 3061 gtggcctttc cactggaagt ggtgcagaag gacagctgct tcaacagccc catgaagctg 3121 cccgtgcaca agctgtacat cgagtacggc atccagcgga tcacatgccc caatgtggac 3181 ggctacttcc ccagcagcgt gaagcctacc atcacctggt acatgggctg ctacaagatc 3241 cagaacttca acaacgtgat ccccgaggga atgaacctga gcttcctgat cgccctgatc 3301 agcaacaacg gcaattacac atgcgtggtc acataccccg agaacggccg gaccttccac 3361 ctgaccagaa cactgacagt gaaggtcgtg ggcagcccca agaatgctgt gcctcctgtg 3421 attcacagcc ccaacgatca cgtggtgtac gagaaagagc ccggcgagga actgctgatc 3481 ccctgtaccg tgtacttcag ctttctgatg gacagccgga acgaagtgtg gtggaccatc 3541 gacggcaaga agcccgacga catcaccatc gatgtgacca tcaacgagag catcagccac 3601 agccggaccg aggacgagac aagaacacag atcctgagca tcaagaaagt gaccagcgag 3661 gacctgaagc ggagctacgt gtgtcatgcc agatccgcca aaggcgaggt ggccaaagcc 3721 gccaaagtga agcagaaggt gcccgctcct cggtacacag tggaactggc ctgtggattc 3781 ggagccacag gcgcgccggg cggcgacaag acccacacct gccccccctg cccagctcca 3841 gaactgctgg gagggccttc cgtgttcctg ttccccccca agcccaagga caccctgatg 3901 atcagcagga cccccgaggt gacctgcgtg gtggtggccg tgtcccacga ggacccagag 3961 gtgaagttca actggtacgt ggacggcgtg gaggtgcaca acgccaagac caagcccaga 4021 gaggagcagt acaacagcac ctacagggtg gtgtccgtgc tgaccgtgct gcaccaggac 4081 tggctgaacg gcaaagaata caagtgcaaa gtctccaaca aggccctggc tgccccaatc 4141 gaaaagacaa tcagcaaggc caagggccag ccacgggagc cccaggtgta caccctgccc 4201 cccagccggg aggagatgac caagaaccag gtgtccctga cctgtctggt gaagggcttc 4261 taccccagcg atatcgccgt ggagtgggag agcaacggcc agcccgagaa caactacaag 4321 accacccccc cagtgctgga cagcgacggc agcttcttcc tgtacagcaa gctgaccgtg 4381 gacaagtcca ggtggcagca gggcaacgtg ttcagctgca gcgtgatgca cgaggccctg 4441 cacaaccact acacccagaa gtccctgagc ctgagccccg gcggcagcgg tcttaatgat 4501 attttcgagg ctcagaagat tgagtggcat gaacaccacc accatcacca ctaaggccgg 4561 cctgtagtca ctggcacaac gcgttacccg gtaagccaat cgggtataca cggtcgtcat 4621 actgcagaca gggttcttct actttgcaag atagtcttga gtagtaaaat aaatagatag 4681 agaaaatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc 4741 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa 4801 attgcatcac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc 4861 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat 4921 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag 4981 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc 5041 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct 5101 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg 5161 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa 5221 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat 5281 atccattttc ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc 5341 atagtataat acgacaaggt gaggaactaa accatgaccg agtacaagcc cacggtgcgc 5401 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac 5461 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg 5521 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac 5581 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc 5641 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg 5701 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc 5761 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag 5821 gcggccgagc gcgccggggt gcccgccttc ctggagacat ccgcgccccg caacctcccc 5881 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc 5941 acctggtgca tgacccgcaa gcccggtgcc tgacacgtgc tacgagattt cgattccacc 6001 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc 6061 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct 6121 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca 6181 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg 6241 tcgacctcta gctagagctt ggcgtaatca tggtcattac caatgcttaa tcagtgaggc 6301 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta 6361 gataactacg atacgggagg gcttaccatc tggccccagc gctgcgatga taccgcgaga 6421 accacgctca ccggctccgg atttatcagc aataaaccag ccagccggaa gggccgagcg 6481 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc 6541 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccatcg ctacaggcat 6601 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag 6661 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat 6721 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa 6781 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa 6841 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga 6901 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg 6961 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc 7021 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg 7081 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatatt 7141 cttcctt //