LOCUS Exported 7236 bp ds-DNA circular SYN 24-MAR-2018 DEFINITION Ligation of SPy_1046-with-NLS-flag into pST1374. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7236) AUTHORS . TITLE Direct Submission JOURNAL Exported Jun 15, 2018 from SnapGene Viewer 4.1.9 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: pST174 Fragment ends: ApaI and XbaI Fragment size: 5108 Insert molecule: SPy_1046-with-NLS-flag Fragment ends: XbaI and ApaI Fragment size: 4114 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..7236 /organism="synthetic DNA construct" /mol_type="other DNA" source 970..1797 /organism="synthetic DNA construct" /mol_type="other DNA" source 1798..1800 /organism="synthetic DNA construct" /mol_type="other DNA" source 1798..1800 /organism="lenti MS2-P65-HSF1_Hygro" /mol_type="other DNA" source 1798..1800 /organism="synthetic DNA construct" /mol_type="other DNA" source 1798..1800 /organism="synthetic DNA construct" /mol_type="other DNA" source 1798..1800 /organism="synthetic DNA construct" /mol_type="other DNA" source 1849..2535 /organism="synthetic DNA construct" /mol_type="other DNA" source 2536..2538 /organism="synthetic DNA construct" /mol_type="other DNA" source 2536..2538 /organism="synthetic DNA construct" /mol_type="other DNA" source 2539..2550 /organism="lenti MS2-P65-HSF1_Hygro" /mol_type="other DNA" source 2539..2550 /organism="synthetic DNA construct" /mol_type="other DNA" source 2800..2832 /organism="lenti dCAS-VP64_Blast" /mol_type="other DNA" source 2833..2835 /organism="synthetic DNA construct" /mol_type="other DNA" source 2839..2841 /organism="synthetic DNA construct" /mol_type="other DNA" source 2839..2841 /organism="synthetic DNA construct" /mol_type="other DNA" source 2839..2841 /organism="lenti MS2-P65-HSF1_Hygro" /mol_type="other DNA" source 2839..2841 /organism="synthetic DNA construct" /mol_type="other DNA" source 2842..2844 /organism="synthetic DNA construct" /mol_type="other DNA" source 2842..2844 /organism="synthetic DNA construct" /mol_type="other DNA" source 2842..2844 /organism="synthetic DNA construct" /mol_type="other DNA" source 2845..3018 /organism="synthetic DNA construct" /mol_type="other DNA" source 3019..3045 /organism="synthetic DNA construct" /mol_type="other DNA" source 3019..3045 /organism="synthetic DNA construct" /mol_type="other DNA" source 3019..3045 /organism="synthetic DNA construct" /mol_type="other DNA" source 3040..3042 /organism="synthetic DNA construct" /mol_type="other DNA" source 3040..3042 /organism="synthetic DNA construct" /mol_type="other DNA" source 3040..3042 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 235..614 /label=CMV enhancer human cytomegalovirus immediate ea /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 615..818 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" promoter 863..881 /label=T7 promoter promoter for bacteriophage T7 RNA p /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 910..930 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 940..963 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 970..1800 /codon_start=1 /label=scFv-GCN4 /label=scFv GCN4 /translation="MGPDIVMTQSPSSLSASVGDRVTITCRSSTGAVTTSNYASWVQEK PGKLFKGLIGGTNNRAPGVPSRFSGSLIGDKATLTISSLQPEDFATYFCALWYSNHWVF GQGTKVELKRGGGGSGGGGSGGGGSSGGGSEVKLLESGGGLVQPGGSLKLSCAVSGFSL TDYGVNWVRQAPGRGLEWIGVIWGDGITDYNSALKDRFIISKDNGKNTVYLQMSKVRSD DTALYYCVTGLFDYWGQGTLVTVSSYPYDVPDYAGGGGGSGGGGSGGGGSGGGGS" CDS 1801..1848 /codon_start=1 /label=XTEN /translation="SGSETPGTSESATPES" CDS 1849..2535 /codon_start=1 /label=APOBEC /translation="MSSETGPVAVDPTLRRRIEPHEFEVFFDPRELRKETCLLYEINWG GRHSIWRHTSQNTNKHVEVNFIEKFTTERYFCPNTRCSITWFLSWSPCGECSRAITEFL SRYPHVTLFIYIARLYHHADPRNRQGLRDLISSGVTIQIMTEQESGYCWRNFVNYSPSN EAHWPRYPHLWVRLYVLELYCIILGLPPCLNILRRKQPQLTFFTIALQSCHYQRLPPHI LWATGLK" misc_feature complement(2025..2044) /label=apobec-seq-rev CDS 2551..2799 /codon_start=1 /label=UGI /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML" CDS 2854..3018 /codon_start=1 /label=GB1 /translation="EYKLILNGKTLKGETTTEAVDAATAEKVFKQYANDNGVDGEWTYD DATKTFTVTE" CDS 3046..3066 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 3100..3141 /codon_start=1 /product="epitope tag from simian virus 5" /label=V5 tag /translation="GKPIPNPLLGLDST" CDS 3151..3168 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 3197..3421 /label=bGH poly /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 3467..3895 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" promoter 3909..4239 /label=SV40 promoter SV40 enhancer and early promoter /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 4090..4225 /label=SV40 ori /note="SV40 origin of replication" promoter 4287..4334 /label=EM7 promoter synthetic bacterial promoter /label=EM7 promoter /note="synthetic bacterial promoter" CDS 4353..4751 /codon_start=1 /gene="Aspergillus terreus BSD" /product="blasticidin S deaminase" /label=Aspergillus terreus BSD /label=BSD /note="confers resistance to blasticidin" /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI KAIVKDSDGQPTAVGIRELLPSGYVWEG" polyA_signal 4909..5030 /label=SV40 poly /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5079..5095) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" protein_bind 5103..5119 /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(5127..5157) /label=lac promoter promoter for the E. coli lac operon /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5172..5193 /label=CAP bindin /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." rep_origin complement(5481..6069) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(6240..7100) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=bla /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(7101..7205) /gene="bla" /label=AmpR promoter ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 atgggtggac 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 accatgggac ctaagaaaaa gaggaaggtg gcggccgctg actacaagga tgacgacgat 961 aaatctagaa tgggtcccga catcgtgatg acccagagcc ccagcagcct gagcgccagc 1021 gtgggcgacc gcgtgaccat cacctgccgc agcagcaccg gcgccgtgac caccagcaac 1081 tacgccagct gggtgcagga gaagcccggc aagctgttca agggcctgat cggcggcacc 1141 aacaaccgcg cccccggcgt gcccagccgc ttcagcggca gcctgatcgg cgacaaggcc 1201 accctgacca tcagcagcct gcagcccgag gacttcgcca cctacttctg cgccctgtgg 1261 tacagcaacc actgggtgtt cggccagggc accaaggtgg agctgaagcg cggcggcggc 1321 ggcagcggcg gcggcggcag cggcggcggc ggcagcagcg gcggcggcag cgaggtgaag 1381 ctgctggaga gcggcggcgg cctggtgcag cccggcggca gcctgaagct gagctgcgcc 1441 gtgagcggct tcagcctgac cgactacggc gtgaactggg tgcgccaggc ccccggccgc 1501 ggcctggagt ggatcggcgt gatctggggc gacggcatca ccgactacaa cagcgccctg 1561 aaggaccgct tcatcatcag caaggacaac ggcaagaaca ccgtgtacct gcagatgagc 1621 aaggtgcgca gcgacgacac cgccctgtac tactgcgtga ccggcctgtt cgactactgg 1681 ggccagggca ccctggtgac cgtgagcagc tacccatacg atgttccaga ttacgctggt 1741 ggaggcggag gttctggggg aggaggtagt ggcggtggtg gttcaggagg cggcggatcc 1801 agcggcagcg agactcccgg gacctcagag tccgccacac ccgaaagtat gagctcagag 1861 actggcccag tggctgtgga ccccacattg agacggcgga tcgagcccca tgagtttgag 1921 gtattcttcg atccgagaga gctccgcaag gagacctgcc tgctttacga aattaattgg 1981 gggggccggc actccatttg gcgacataca tcacagaaca ctaacaagca cgtcgaagtc 2041 aacttcatcg agaagttcac gacagaaaga tatttctgtc cgaacacaag gtgcagcatt 2101 acctggtttc tcagctggag cccatgcggc gaatgtagta gggccatcac tgaattcctg 2161 tcaaggtatc cccacgtcac tctgtttatt tacatcgcaa ggctgtacca ccacgctgac 2221 ccccgcaatc gacaaggcct gcgggatttg atctcttcag gtgtgactat ccaaattatg 2281 actgagcagg agtcaggata ctgctggaga aactttgtga attatagccc gagtaatgaa 2341 gcccactggc ctaggtatcc ccatctgtgg gtacgactgt acgttcttga actgtactgc 2401 atcatactgg gcctgcctcc ttgtctcaac attctgagaa ggaagcagcc acagctgaca 2461 ttctttacca tcgctcttca gtcttgtcat taccagcgac tgcccccaca cattctctgg 2521 gccaccgggt tgaaaggcgg aggtggaagc actaatctgt cagatattat tgaaaaggag 2581 accggaaagc aactggttat ccaggaatcc atcctcatgc tcccagagga ggtggaagaa 2641 gtcattggga acaagccgga aagcgatata ctcgtgcaca ccgcctacga cgagagcacc 2701 gacgagaatg tcatgcttct gactagcgac gcccctgaat acaagccttg ggctctggtc 2761 atacaggata gcaacggtga gaacaagatt aagatgctcg gaggaggagg aagcggagga 2821 ggaggtagcg gaggaggtgg aagccggacc gaagagtaca agcttatcct gaacggtaaa 2881 accctgaaag gtgaaaccac caccgaagct gttgacgctg ctaccgcgga aaaagttttc 2941 aaacagtacg ctaacgacaa cggtgttgac ggtgaatgga cctacgacga cgctaccaaa 3001 accttcacgg taaccgaagg tggtggtagc ggtggtggtg gtagtcccaa gaagaagagg 3061 aaagtctcga gcggtggagc tgcaggaggg cccttcgaag gtaagcctat ccctaaccct 3121 ctcctcggtc tcgattctac gcgtaccggt catcatcacc atcaccattg agtttaaacc 3181 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc 3241 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa 3301 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac 3361 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg 3421 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc 3481 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc 3541 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt 3601 ccccgtcaag ctctaaatcg gggcatccct ttagggttcc gatttagtgc tttacggcac 3661 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag 3721 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa 3781 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgggg 3841 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc 3901 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccaggc aggcagaagt 3961 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca 4021 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta 4081 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga 4141 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag 4201 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata 4261 tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag tatatcggca 4321 tagtataata cgacaaggtg aggaactaaa ccatggccaa gcctttgtct caagaagaat 4381 ccaccctcat tgaaagagca acggctacaa tcaacagcat ccccatctct gaagactaca 4441 gcgtcgccag cgcagctctc tctagcgacg gccgcatctt cactggtgtc aatgtatatc 4501 attttactgg gggaccttgt gcagaactcg tggtgctggg cactgctgct gctgcggcag 4561 ctggcaacct gacttgtatc gtcgcgatcg gaaatgagaa caggggcatc ttgagcccct 4621 gcggacggtg tcgacaggtg cttctcgatc tgcatcctgg gatcaaagcg atagtgaagg 4681 acagtgatgg acagccgacg gcagttggga ttcgtgaatt gctgccctct ggttatgtgt 4741 gggagggcta agcacttcgt ggccgaggag caggactgac acgtgctacg agatttcgat 4801 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg 4861 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt 4921 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt 4981 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt 5041 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga 5101 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc 5161 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc 5221 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc 5281 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt 5341 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca 5401 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa 5461 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat 5521 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc 5581 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc 5641 gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt 5701 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac 5761 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg 5821 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca 5881 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc 5941 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa 6001 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa 6061 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac 6121 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta 6181 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt 6241 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata 6301 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc 6361 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac 6421 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag 6481 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac 6541 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc 6601 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg 6661 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc 6721 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct 6781 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc 6841 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc 6901 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc 6961 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc 7021 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca 7081 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt 7141 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt 7201 ccgcgcacat ttccccgaaa agtgccacct gacgtc //