LOCUS Exported 7846 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-SEMA6C-Fc(DAPA)-AviTag-6xHis SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7846) 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 7846) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7846 /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 5188..5232 /codon_start=1 /product="peptide tag that allows for enzymatic biotinylation" /label=AviTag(TM) /translation="GLNDIFEAQKIEWHE" CDS 5233..5250 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(5390..5407) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 5396..5507 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(5625..5645) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 5630..5959 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 5810..5945 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5872..5891 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 6007..6054 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 6073..6672 /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(6073..6092) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 6569..6589 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 6802..6923 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(6839..6858) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 6893..6912 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" CDS complement(6976..7836) /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 7599..7618 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7837..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 cggccgccgc ttttccacaa 2761 gatcctctgc ctctgctgat tagcgacctg cagggaacaa gccctctgag ctggttcaga 2821 ggcctggaag atgatgccgt ggccgccgaa ctgggcctcg attttcagcg gtttctgacc 2881 ctgaacagaa ccctgctggt ggccgccaga gatcacgtgt tcagcttcga tctgcaagcc 2941 gaggaagaag gcgaaggact ggtccccaac aagtacctga cttggagaag ccaggacgtg 3001 gaaaattgtg ccgtgcgggg caagctgacc gacgagtgct acaactacat cagagtgctg 3061 gtgccctggg actctcagac actgctggcc tgtggcacca atagcttcag ccctgtgtgc 3121 agaagctacg gcatcaccag tctgcagcaa gagggcgaag aactgagcgg acaggccaga 3181 tgtcctttcg acgccacaca gagcaatgtg gccatctttg ccgagggcag cctgtattct 3241 gccacagccg ctgattttca ggccagcgac gccgtggtgt atagatctct tggaccccag 3301 cctcctctgc ggagcgccaa atatgatagc aagtggctgc gcgagcctca cttcgtgcaa 3361 gctctggaac acggcgacca cgtgtacttt ttctttagag aggtgtccgt cgaggacgcc 3421 agactgggca gagtgcagtt tagcagagtg gccagagtgt gcaagagaga catgggcgga 3481 agccccagag cactggatag acactggacc agctttctga agctgcggct gaactgtagc 3541 gtgcccggcg atagcacctt ctactttgat gtgctgcagg ccctgacagg ccctgtgaat 3601 ctgcatggaa gatctgccct gttcggcgtg ttcaccacac agacaaatag catccctggc 3661 agcgccgtgt gcgccttcta tctggatgag atcgagagag gcttcgaggg caagttcaaa 3721 gagcagagaa gtctggacgg cgcctggaca cctgtgtctg aggatagagt gccatctcct 3781 agacctggct cttgtgctgg tgttggcgga gccgctctgt tcagctctag cagagatctg 3841 cctgacgacg tgctgacctt catcaaggct catcctctgc tggaccctgc cgtgcctcct 3901 gttacacatc aaccactgct gactctgacc agcagagccc tgctgacaca ggttgcagtg 3961 gatggaatgg ccggacctca cagcaatatc accgtgatgt ttctgggcag caacgacggc 4021 acagtgctga aagttctgac acctggcgga agaagcggag gccctgaacc tatcctgctg 4081 gaagagattg acgcctactc tcctgccaga tgcagcggca aaagaacagc ccagaccgcc 4141 agaagaatca tcggcctgga actggatacc gagggccaca gactgtttgt ggcctttagc 4201 ggctgcatcg tgtacctgcc tctgagcaga tgtgccagac atggcgcctg tcagagaagc 4261 tgtctggcct ctcaggaccc ttactgtgga tggcacagct ccagaggctg tgtggatatt 4321 agaggcagcg gcggcacaga tgtggaccag gccggaaatc aagagtccat ggaacatggc 4381 gattgccagg atggcgccac aggatctcaa tctggacctg gcgattctgc ctacggcgtc 4441 agaagagatt tgcctccagc cagcgcaagc agaagcgtgc caattcctgg cgcgccgggc 4501 ggcgacaaga cccacacctg ccccccctgc ccagctccag aactgctggg agggccttcc 4561 gtgttcctgt tcccccccaa gcccaaggac accctgatga tcagcaggac ccccgaggtg 4621 acctgcgtgg tggtggccgt gtcccacgag gacccagagg tgaagttcaa ctggtacgtg 4681 gacggcgtgg aggtgcacaa cgccaagacc aagcccagag aggagcagta caacagcacc 4741 tacagggtgg tgtccgtgct gaccgtgctg caccaggact ggctgaacgg caaagaatac 4801 aagtgcaaag tctccaacaa ggccctggct gccccaatcg aaaagacaat cagcaaggcc 4861 aagggccagc cacgggagcc ccaggtgtac accctgcccc ccagccggga ggagatgacc 4921 aagaaccagg tgtccctgac ctgtctggtg aagggcttct accccagcga tatcgccgtg 4981 gagtgggaga gcaacggcca gcccgagaac aactacaaga ccaccccccc agtgctggac 5041 agcgacggca gcttcttcct gtacagcaag ctgaccgtgg acaagtccag gtggcagcag 5101 ggcaacgtgt tcagctgcag cgtgatgcac gaggccctgc acaaccacta cacccagaag 5161 tccctgagcc tgagccccgg cggcagcggt cttaatgata ttttcgaggc tcagaagatt 5221 gagtggcatg aacaccacca ccatcaccac taaggccggc ctgtagtcac tggcacaacg 5281 cgttacccgg taagccaatc gggtatacac ggtcgtcata ctgcagacag ggttcttcta 5341 ctttgcaaga tagtcttgag tagtaaaata aatagataga gaaaatcagc ctcgactgtg 5401 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa 5461 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcaca acactcaacc 5521 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa 5581 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt 5641 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa 5701 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag 5761 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct 5821 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc 5881 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg 5941 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca 6001 gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg 6061 aggaactaaa ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc 6121 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc 6181 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc 6241 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg 6301 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc 6361 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac 6421 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc 6481 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg 6541 cccgccttcc tggagacatc cgcgccccgc aacctcccct tctacgagcg gctcggcttc 6601 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag 6661 cccggtgcct gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg 6721 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg 6781 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc 6841 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg 6901 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg 6961 gcgtaatcat ggtcattacc aatgcttaat cagtgaggca cctatctcag cgatctgtct 7021 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg 7081 cttaccatct ggccccagcg ctgcgatgat accgcgagaa ccacgctcac cggctccgga 7141 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt 7201 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt 7261 taatagtttg cgcaacgttg ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt 7321 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat 7381 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc 7441 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc 7501 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat 7561 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag 7621 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt 7681 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc 7741 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa 7801 gggaataagg gcgacacgga aatgttgaat actcatattc ttcctt //