LOCUS Exported 8347 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-CD1A-COMP5AP-AviTag-9xHis SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8347) 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 8347) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8347 /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 3997..4020 /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /label=HRV 3C site /translation="LEVLFQGP" CDS 5680..5724 /codon_start=1 /product="peptide tag that allows for enzymatic biotinylation" /label=AviTag(TM) /translation="GLNDIFEAQKIEWHE" CDS 5725..5751 /codon_start=1 /product="9xHis affinity tag" /label=9xHis /translation="HHHHHHHHH" primer_bind complement(5891..5908) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 5897..6008 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(6126..6146) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 6131..6460 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 6311..6446 /label=SV40 ori /note="SV40 origin of replication" primer_bind 6373..6392 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 6508..6555 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 6574..7173 /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(6574..6593) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 7070..7090 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 7303..7424 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(7340..7359) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 7394..7413 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" CDS complement(7477..8337) /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 8100..8119 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(8338..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 cggccgccat tcagagaacc 2761 cctaagatcc aggtgtacag cagacacccc gccgagaacg gcaagagcaa cttcctgaac 2821 tgctacgtgt ccggctttca ccccagcgac attgaggtgg acctgctgaa gaacggcgag 2881 cggatcgaga aggtggaaca cagcgatctg agcttcagca aggactggtc cttctacctg 2941 ctgtactaca ccgagttcac ccctaccgag aaggacgagt acgcctgcag agtgaaccac 3001 gtgacactga gccagcctaa gatcgtgaag tgggacagag acatgggcgg aggcggatct 3061 ggtggcggag gaagtggcgg cggaggatct aatgccgatg gcctgaaaga gcccctgagc 3121 ttccacgtga cctggatcgc cagcttctac aaccacagct ggaagcagaa cctggtgtct 3181 ggctggctga gcgatctgca gacccacacc tgggacagca acagcagcac catcgtgttt 3241 ctgtgcccct ggtccagagg caacttcagc aacgaggaat ggaaagagct ggaaaccctg 3301 ttccggatca gaaccatcag aagcttcgag ggcatcagaa gatacgccca cgagctgcag 3361 ttcgagtacc ccttcgagat ccaagtgaca ggcggatgcg agctgcactc tggcaaagtg 3421 tctggcagct ttctgcagct ggcctaccag ggcagcgact tcgtgtcctt ccagaacaac 3481 agctggctgc cctatcctgt ggccggcaat atggccaagc acttctgcaa ggtgctgaac 3541 cagaaccagc acgagaacga catcacccac aacctgctga gcgacacatg ccccagattc 3601 atcctgggac tgctggatgc cggaaaggcc catctgcaga gacaagtgaa gcctgaggcc 3661 tggctgtctc acggaccttc tccaggacct ggacatctgc agcttgtgtg ccatgtgtcc 3721 ggattctacc ccaagcctgt gtgggtcatg tggatgagag gcgagcaaga gcagcagggc 3781 acacagagag gcgatatcct gcctagcgcc gacggcacct ggtatctgag agctacactg 3841 gaagtggccg ctggcgaagc cgctgatctg tcctgtagag tgaagcacag cagcctggaa 3901 ggccaggaca tcgtgctgta ttgggagcac cacagcagcg ttggcgcgcc ggaagatcaa 3961 gttgatccaa gattgattga tggcaagggc ggatctctgg aggtgctgtt ccagggacca 4021 ggcggaagcg gcaatcccgg cggcgatctg gccccccaga tgctgcgcga gctgcaggag 4081 accaatgccg ccctgcagga cgtgcgcgag ctgctgcgcc accgcgtgaa ggagatcacc 4141 ttcctgaaga acaccgtgat ggagtgcgac gcctgcggta tgcagccctc tagaggctcc 4201 ggctccatca tcccagttga ggaggagaac ccggacttct ggaaccgcga ggcagccgag 4261 gccctgggtg ccgccaagaa gctgcagcct gcacagacag ccgccaagaa cctcatcatc 4321 ttcctgggcg atgggatggg ggtgtctacg gtgacagctg ccaggatcct aaaagggcag 4381 aagaaggaca aactggggcc tgagataccc ctggccatgg accgcttccc atatgtggct 4441 ctgtccaaga catacaatgt agacaaacat gtgccagaca gtggagccac agccacggcc 4501 tacctgtgcg gggtcaaggg caacttccag accattggct tgagtgcagc cgcccgcttt 4561 aaccagtgca acacgacacg cggcaacgag gtcatctccg tgatgaatcg ggccaagaaa 4621 gcagggaagt cagtgggagt ggtaaccacc acacgagtgc agcacgcctc gccagccggc 4681 acctacgccc acacggtgaa ccgcaactgg tactcggacg ccgacgtgcc tgcctcggcc 4741 cgccaggagg ggtgccagga catcgctacg cagctcatct ccaacatgga cattgacgtg 4801 atcctaggtg gaggccgaaa gtacatgttt cgcatgggaa ccccagaccc tgagtaccca 4861 gatgactaca gccaaggtgg gaccaggctg gacgggaaga atctggtgca ggaatggctg 4921 gcgaagcgcc agggtgcccg gtatgtgtgg aaccgcactg agctcatgca ggcttccctg 4981 gacccgtctg tgacccatct catgggtctc tttgagcctg gagacatgaa atacgagatc 5041 caccgagact ccacactgga cccctccctg atggagatga cagaggctgc cctgcgcctg 5101 ctgagcagga acccccgcgg cttcttcctc ttcgtggagg gtggtcgcat cgaccatggt 5161 catcatgaaa gcagggctta ccgggcactg actgagacga tcatgttcga cgacgccatt 5221 gagagggcgg gccagctcac cagcgaggag gacacgctga gcctcgtcac tgccgaccac 5281 tcccacgtct tctccttcgg aggctacccc ctgcgaggga gctccatctt cgggctggcc 5341 cctggcaagg cccgggacag gaaggcctac acggtcctcc tatacggaaa cggtccaggc 5401 tatgtgctca aggacggcgc ccggccggat gttaccgaga gcgagagcgg gagccccgag 5461 tatcggcagc agtcagcagt gcccctggac gaagagaccc acgcaggcga ggacgtggcg 5521 gtgttcgcgc gcggcccgca ggcgcacctg gttcacggcg tgcaggagca gaccttcata 5581 gcgcacgtca tggccttcgc cgcctgcctg gagccctaca ccgcctgcga cctggcgccc 5641 cccgccggca ccaccgacgc cgcgcacccg ggtggcggag gactgaacga catcttcgag 5701 gctcagaaaa tcgaatggca cgaacatcac catcaccatc accatcacca ttaaggccgg 5761 cctgtagtca ctggcacaac gcgttacccg gtaagccaat cgggtataca cggtcgtcat 5821 actgcagaca gggttcttct actttgcaag atagtcttga gtagtaaaat aaatagatag 5881 agaaaatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc 5941 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa 6001 attgcatcac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc 6061 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat 6121 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag 6181 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc 6241 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct 6301 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg 6361 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa 6421 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat 6481 atccattttc ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc 6541 atagtataat acgacaaggt gaggaactaa accatgaccg agtacaagcc cacggtgcgc 6601 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac 6661 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg 6721 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac 6781 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc 6841 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg 6901 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc 6961 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag 7021 gcggccgagc gcgccggggt gcccgccttc ctggagacat ccgcgccccg caacctcccc 7081 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc 7141 acctggtgca tgacccgcaa gcccggtgcc tgacacgtgc tacgagattt cgattccacc 7201 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc 7261 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct 7321 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca 7381 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg 7441 tcgacctcta gctagagctt ggcgtaatca tggtcattac caatgcttaa tcagtgaggc 7501 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta 7561 gataactacg atacgggagg gcttaccatc tggccccagc gctgcgatga taccgcgaga 7621 accacgctca ccggctccgg atttatcagc aataaaccag ccagccggaa gggccgagcg 7681 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc 7741 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccatcg ctacaggcat 7801 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag 7861 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat 7921 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa 7981 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa 8041 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga 8101 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg 8161 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc 8221 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg 8281 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatatt 8341 cttcctt //