LOCUS Exported 6379 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for superfolder GFP-fused human chemokine CXCL12. ACCESSION . VERSION . KEYWORDS pcDNA3-CXCL12-sfGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6379) AUTHORS Heredia JD, Park J, Brubaker RJ, Szymanski SK, Gill KS, Procko E TITLE Mapping Interaction Sites on Human Chemokine Receptors by Deep Mutational Scanning. JOURNAL J Immunol. 2018 Apr 20. pii: jimmunol.1800343. doi: 10.4049/jimmunol.1800343. PUBMED 29678950 REFERENCE 2 (bases 1 to 6379) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6379 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(44..63) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 235..614 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 615..818 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 769..789 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 863..882 /label=T7 /note="T7 promoter, forward primer" promoter 863..881 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 1219..1932 /codon_start=1 /product="GFP variant that folds robustly even when fused to poorly folded proteins (Pedelacq et al., 2006)" /label=superfolder GFP /note="mammalian codon-optimized" /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF VTAAGITHGMDELYK" primer_bind complement(1246..1274) /label=GFP-R /note="GFP, reverse primer. Does NOT anneal to EGFP" primer_bind 1869..1889 /label=GFP-F /note="GFP, forward primer. Does NOT anneal to EGFP" primer_bind complement(1973..1990) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 1979..2203 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 2249..2677 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2336..2355) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2546..2567 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(2686..2706) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 2691..3020 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 2871..3006 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2933..2952 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3087..3881 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3141..3160) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3751..3770 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 4055..4176 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4092..4111) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4146..4165 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(4225..4241) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4225..4241) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4238..4260) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4249..4265 /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)." promoter complement(4273..4303) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4318..4339 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(4456..4473) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4627..5212) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4707..4726) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5383..6243) /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 6006..6025 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6244..6348) /gene="bla" /label=AmpR promoter ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc 901 gccaccatga acgccaaggt cgtggtcgtg ctggtcctcg tgctgaccgc gctctgcctc 961 agcgacggga agcccgtcag cctgagctac agatgcccat gccgattctt cgaaagccat 1021 gttgccagag ccaacgtcaa gcatctcaaa attctcaaca ctccaaactg tgcccttcag 1081 attgtagccc ggctgaagaa caacaacaga caagtgtgca ttgacccgaa gctaaagtgg 1141 attcaggagt acctggagaa agctttaaac aagagcggcg gagggggctc atctggaggt 1201 gggtccggta gtggtggatc caaaggagaa gaacttttca ctggagttgt cccaattctt 1261 gttgaattag atggtgatgt taatgggcac aaattttctg tccgtggaga gggtgaaggt 1321 gatgctacaa acggaaaact cacccttaaa tttatttgca ctactggaaa actacctgtt 1381 ccatggccaa cacttgtcac tactctgacc tatggtgttc aatgcttttc ccgttatccg 1441 gatcacatga aacggcatga ctttttcaag agtgccatgc ccgaaggtta tgtacaggaa 1501 cgcactatat ctttcaaaga tgacgggacc tacaagacgc gtgctgaagt caagtttgaa 1561 ggtgataccc ttgttaatcg tatcgagtta aaaggtattg attttaaaga agatggaaac 1621 attctcggac acaaactgga gtacaacttt aactcacaca atgtatacat cacggcagac 1681 aaacaaaaga atggaatcaa agctaacttc aaaattcgcc acaacgttga agatggttcc 1741 gttcaactag cagaccatta tcaacaaaat actccaattg gcgatggccc tgtcctttta 1801 ccagacaacc attacctgtc gacacaatct gtcctttcga aagatcccaa cgaaaagcgt 1861 gaccacatgg tccttcttga gtttgtaact gctgctggga ttacacatgg catggatgag 1921 ctctacaaat gataactcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact 1981 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg 2041 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg 2101 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg 2161 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga 2221 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg 2281 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct 2341 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat 2401 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt 2461 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg 2521 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac 2581 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta 2641 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt 2701 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca 2761 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa 2821 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc 2881 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg 2941 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg 3001 gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc 3061 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc 3121 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct 3181 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg 3241 acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca 3301 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc 3361 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga 3421 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc 3481 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc 3541 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg 3601 ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct 3661 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc 3721 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc 3781 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc 3841 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga 3901 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt 3961 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg 4021 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg 4081 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc 4141 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc 4201 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct 4261 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg 4321 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct 4381 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg 4441 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc 4501 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg 4561 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct 4621 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca 4681 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct 4741 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc 4801 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt 4861 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc 4921 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc 4981 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg 5041 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc 5101 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag 5161 cggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc 5221 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt 5281 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt 5341 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag 5401 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt 5461 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc 5521 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc 5581 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg 5641 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac 5701 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg 5761 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc 5821 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact 5881 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc 5941 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat 6001 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc 6061 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac 6121 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa 6181 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact 6241 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg 6301 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg 6361 aaaagtgcca cctgacgtc //