LOCUS Exported 6819 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Gateway entry vector with EF1-a promoter, V5-His C-term, Lac Z. ACCESSION . VERSION . KEYWORDS pEF-ENTR-LacZ (710-1) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6819) AUTHORS Campeau E, Ruhl VE, Rodier F, Smith CL, Rahmberg BL, Fuss JO, Campisi J, Yaswen P, Cooper PK, Kaufman PD. TITLE A versatile viral system for expression and depletion of proteins in mammalian cells. JOURNAL PLoS One. 2009 Aug 6;4(8):e6529. PUBMED 19657394 REFERENCE 2 (bases 1 to 6819) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6819 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 103..189 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" terminator 281..308 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" primer_bind 316..339 /label=pENTR-F /note="pENTR vectors, forward primer" protein_bind 358..457 /gene="mutant version of attL" /label=attL1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 588..1766 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 819..1757 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 1714..1734 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" primer_bind 1783..1802 /label=T7 /note="T7 promoter, forward primer" promoter 1783..1801 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 1895..4939 /codon_start=1 /gene="lacZ" /product="beta-galactosidase" /label=lacZ /translation="VVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRS LNGEWRFAWFPVPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNVTYPITVNPP FVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVNSAFHLWCNGRWVGYGQDSRLPSEF DLSAFLRAGENRLAVMVLRWSDGSYLEDQDMWRMSGIFRDVSLLHKPTTQISDFHVATR FNDDFSRAVLEAEVQMCGELRDYLRVTVSLWQGETQVASGTAPFGGEIIDERGGYADRV TLRLNVENPKLWSAEIPNLYRAVVELHTADGTLIEAEACDVGFREVRIENGLLLLNGKP LLIRGVNRHEHHPLHGQVMDEQTMVQDILLMKQNNFNAVRCSHYPNHPLWYTLCDRYGL YVVDEANIETHGMVPMNRLTDDPRWLPAMSERVTRMVQRDRNHPSVIIWSLGNESGHGA NHDALYRWIKSVDPSRPVQYEGGGADTTATDIICPMYARVDEDQPFPAVPKWSIKKWLS LPGETRPLILCEYAHAMGNSLGGFAKYWQAFRQYPRLQGGFVWDWVDQSLIKYDENGNP WSAYGGDFGDTPNDRQFCMNGLVFADRTPHPALTEAKHQQQFFQFRLSGQTIEVTSEYL FRHSDNELLHWMVALDGKPLASGEVPLDVAPQGKQLIELPELPQPESAGQLWLTVRVVQ PNATAWSEAGHISAWQQWRLAENLSVTLPAASHAIPHLTTSEMDFCIELGNKRWQFNRQ SGFLSQMWIGDKKQLLTPLRDQFTRAPLDNDIGVSEATRIDPNAWVERWKAAGHYQAEA ALLQCTADTLADAVLITTAHAWQHQGKTLFISRKTYRIDGSGQMAITVDVEVASDTPHP ARIGLNCQLAQVAERVNWLGLGPQENYPDRLTAACFDRWDLPLSDMYTPYVFPSENGLR CGTRELNYGPHQWRGDFQFNISRYSQQQLMETSHRHLLHAEEGTWLNIDGFHMGIGGDD SWSPSVSAEFQLSAGRYHYQLVWCQK" primer_bind complement(1897..1919) /label=M13/pUC Forward /note="In lacZ gene" CDS 4967..5008 /codon_start=1 /product="epitope tag from simian virus 5" /label=V5 tag /translation="GKPIPNPLLGLDST" CDS 5018..5035 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" protein_bind complement(5049..5148) /gene="mutant version of attL" /label=attL2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" primer_bind complement(5260..5279) /label=pENTR-R /note="pENTR vectors, reverse primer" CDS 5271..6080 /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPGDAWLLTTAIPGKTA FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind complement(5338..5357) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" rep_origin 6170..6758 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6659..6678 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgctagca tggatctcgg 61 ggacgtctaa ctactaagcg agagtaggga actgccaggc atcaaataaa acgaaaggct 121 cagtcggaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt 181 aggacaaatc cgccgggagc ggatttgaac gttgtgaagc aacggcccgg agggtggcgg 241 gcaggacgcc cgccataaac tgccaggcat caaactaagc agaaggccat cctgacggat 301 ggcctttttg cgtttctaca aactcttcct gttagttagt tacttaagct cgggccccaa 361 ataatgattt tattttgact gatagtgacc tgttcgttgc aacaaattga taagcaatgc 421 ttttttataa tgccaacttt gtacaaaaaa gcaggctcca ccatgggaac cgacattgat 481 tattgactag gcttttgcaa aaagctttgc aaagatggat aaagttttaa acagagagga 541 atctttgcag ctaatggacc ttctaggtct tgaaaggagt gggaattggc tccggtgccc 601 gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga ggggtcggca 661 attgaaccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact 721 ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta gtcgccgtga 781 acgttctttt tcgcaacggg tttgccgcca gaacacaggt aagtgccgtg tgtggttccc 841 gcgggcctgg cctctttacg ggttatggcc cttgcgtgcc ttgaattact tccacctggc 901 tgcagtacgt gattcttgat cccgagcttc gggttggaag tgggtgggag agttcgaggc 961 cttgcgctta aggagcccct tcgcctcgtg cttgagttga ggcctggcct gggcgctggg 1021 gccgccgcgt gcgaatctgg tggcaccttc gcgcctgtct cgctgctttc gataagtctc 1081 tagccattta aaatttttga tgacctgctg cgacgctttt tttctggcaa gatagtcttg 1141 taaatgcggg ccaagatctg cacactggta tttcggtttt tggggccgcg ggcggcgacg 1201 gggcccgtgc gtcccagcgc acatgttcgg cgaggcgggg cctgcgagcg cggccaccga 1261 gaatcggacg ggggtagtct caagctggcc ggcctgctct ggtgcctggc ctcgcgccgc 1321 cgtgtatcgc cccgccctgg gcggcaaggc tggcccggtc ggcaccagtt gcgtgagcgg 1381 aaagatggcc gcttcccggc cctgctgcag ggagctcaaa atggaggacg cggcgctcgg 1441 gagagcgggc gggtgagtca cccacacaaa ggaaaagggc ctttccgtcc tcagccgtcg 1501 cttcatgtga ctccacggag taccgggcgc cgtccaggca cctcgattag ttctcgagct 1561 tttggagtac gtcgtcttta ggttgggggg aggggtttta tgcgatggag tttccccaca 1621 ctgagtgggt ggagactgaa gttaggccag cttggcactt gatgtaattc tccttggaat 1681 ttgccctttt tgagtttgga tcttggttca ttctcaagcc tcagacagtg gttcaaagtt 1741 tttttcttcc atttcaggtg tcgtgaggaa ttagcttggt actaatacga ctcactatag 1801 ggagacccaa gctggctagg taagcttggt accgagctcg gatccactag tccagtgtgg 1861 tggaattctg cagatcgaaa cgatgataga tcccgtcgtt ttacaacgtc gtgactggga 1921 aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg 1981 taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga 2041 atggcgcttt gcctggtttc cggtaccaga agcggtgccg gaaagctggc tggagtgcga 2101 tcttcctgag gccgatactg tcgtcgtccc ctcaaactgg cagatgcacg gttacgatgc 2161 gcccatctac accaacgtaa cctatcccat tacggtcaat ccgccgtttg ttcccacgga 2221 gaatccgacg ggttgttact cgctcacatt taatgttgat gaaagctggc tacaggaagg 2281 ccagacgcga attatttttg atggcgttaa ctcggcgttt catctgtggt gcaacgggcg 2341 ctgggtcggt tacggccagg acagtcgttt gccgtctgaa tttgacctga gcgcattttt 2401 acgcgccgga gaaaaccgcc tcgcggtgat ggtgctgcgt tggagtgacg gcagttatct 2461 ggaagatcag gatatgtggc ggatgagcgg cattttccgt gacgtctcgt tgctgcataa 2521 accgactaca caaatcagcg atttccatgt tgccactcgc tttaatgatg atttcagccg 2581 cgctgtactg gaggctgaag ttcagatgtg cggcgagttg cgtgactacc tacgggtaac 2641 agtttcttta tggcagggtg aaacgcaggt cgccagcggc accgcgcctt tcggcggtga 2701 aattatcgat gagcgtggtg gttatgccga tcgcgtcaca ctacgtctga acgtcgaaaa 2761 cccgaaactg tggagcgccg aaatcccgaa tctctatcgt gcggtggttg aactgcacac 2821 cgccgacggc acgctgattg aagcagaagc ctgcgatgtc ggtttccgcg aggtgcggat 2881 tgaaaatggt ctgctgctgc tgaacggcaa gccgttgctg attcgaggcg ttaaccgtca 2941 cgagcatcat cctctgcatg gtcaggtcat ggatgagcag acgatggtgc aggatatcct 3001 gctgatgaag cagaacaact ttaacgccgt gcgctgttcg cattatccga accatccgct 3061 gtggtacacg ctgtgcgacc gctacggcct gtatgtggtg gatgaagcca atattgaaac 3121 ccacggcatg gtgccaatga atcgtctgac cgatgatccg cgctggctac cggcgatgag 3181 cgaacgcgta acgcgaatgg tgcagcgcga tcgtaatcac ccgagtgtga tcatctggtc 3241 gctggggaat gaatcaggcc acggcgctaa tcacgacgcg ctgtatcgct ggatcaaatc 3301 tgtcgatcct tcccgcccgg tgcagtatga aggcggcgga gccgacacca cggccaccga 3361 tattatttgc ccgatgtacg cgcgcgtgga tgaagaccag cccttcccgg ctgtgccgaa 3421 atggtccatc aaaaaatggc tttcgctacc tggagagacg cgcccgctga tcctttgcga 3481 atacgcccac gcgatgggta acagtcttgg cggtttcgct aaatactggc aggcgtttcg 3541 tcagtatccc cgtttacagg gcggcttcgt ctgggactgg gtggatcagt cgctgattaa 3601 atatgatgaa aacggcaacc cgtggtcggc ttacggcggt gattttggcg atacgccgaa 3661 cgatcgccag ttctgtatga acggtctggt ctttgccgac cgcacgccgc atccagcgct 3721 gacggaagca aaacaccagc agcagttttt ccagttccgt ttatccgggc aaaccatcga 3781 agtgaccagc gaatacctgt tccgtcatag cgataacgag ctcctgcact ggatggtggc 3841 gctggatggt aagccgctgg caagcggtga agtgcctctg gatgtcgctc cacaaggtaa 3901 acagttgatt gaactgcctg aactaccgca gccggagagc gccgggcaac tctggctcac 3961 agtacgcgta gtgcaaccga acgcgaccgc atggtcagaa gccgggcaca tcagcgcctg 4021 gcagcagtgg cgtctggcgg aaaacctcag tgtgacgctc cccgccgcgt cccacgccat 4081 cccgcatctg accaccagcg aaatggattt ttgcatcgag ctgggtaata agcgttggca 4141 atttaaccgc cagtcaggct ttctttcaca gatgtggatt ggcgataaaa aacaactgct 4201 gacgccgctg cgcgatcagt tcacccgtgc accgctggat aacgacattg gcgtaagtga 4261 agcgacccgc attgacccta acgcctgggt cgaacgctgg aaggcggcgg gccattacca 4321 ggccgaagca gcgttgttgc agtgcacggc agatacactt gctgatgcgg tgctgattac 4381 gaccgctcac gcgtggcagc atcaggggaa aaccttattt atcagccgga aaacctaccg 4441 gattgatggt agtggtcaaa tggcgattac cgttgatgtt gaagtggcga gcgatacacc 4501 gcatccggcg cggattggcc tgaactgcca gctggcgcag gtagcagagc gggtaaactg 4561 gctcggatta gggccgcaag aaaactatcc cgaccgcctt actgccgcct gttttgaccg 4621 ctgggatctg ccattgtcag acatgtatac cccgtacgtc ttcccgagcg aaaacggtct 4681 gcgctgcggg acgcgcgaat tgaattatgg cccacaccag tggcgcggcg acttccagtt 4741 caacatcagc cgctacagtc aacagcaact gatggaaacc agccatcgcc atctgctgca 4801 cgcggaagaa ggcacatggc tgaatatcga cggtttccat atggggattg gtggcgacga 4861 ctcctggagc ccgtcagtat cggcggaatt ccagctgagc gccggtcgct accattacca 4921 gttggtctgg tgtcaaaaag cggccgctcg aggtcaccca ttcgaaggta agcctatccc 4981 taaccctctc ctcggtctcg attctacgcg taccggtcat catcaccatc accattgagt 5041 ttatctagac ccagctttct tgtacaaagt tggcattata agaaagcatt gcttatcaat 5101 ttgttgcaac gaacaggtca ctatcagtca aaataaaatc attatttgcc atccagctgc 5161 agctctggcc cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat 5221 catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata 5281 ttcaacggga aacgtcgagg ccgcgattaa attccaacat ggatgctgat ttatatgggt 5341 ataaatgggc tcgcgataat gtcgggcaat caggtgcgac aatctatcgc ttgtatggga 5401 agcccgatgc gccagagttg tttctgaaac atggcaaagg tagcgttgcc aatgatgtta 5461 cagatgagat ggtcagacta aactggctga cggaatttat gcctcttccg accatcaagc 5521 attttatccg tactcctggt gatgcatggt tactcaccac tgcgatcccc ggaaaaacag 5581 cattccaggt attagaagaa tatcctgatt caggtgaaaa tattgttgat gcgctggcag 5641 tgttcctgcg ccggttgcat tcgattcctg tttgtaattg tccttttaac agcgatcgcg 5701 tatttcgtct cgctcaggcg caatcacgaa tgaataacgg tttggttgat gcgagtgatt 5761 ttgatgacga gcgtaatggc tggcctgttg aacaagtctg gaaagaaatg cataaacttt 5821 tgccattctc accggattca gtcgtcactc atggtgattt ctcacttgat aaccttattt 5881 ttgacgaggg gaaattaata ggttgtattg atgttggacg agtcggaatc gcagaccgat 5941 accaggatct tgccatccta tggaactgcc tcggtgagtt ttctccttca ttacagaaac 6001 ggctttttca aaaatatggt attgataatc ctgatatgaa taaattgcag tttcatttga 6061 tgctcgatga gtttttctaa tcagaattgg ttaattggtt gtaacattat tcagattggg 6121 ccccgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt 6181 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt 6241 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc 6301 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg 6361 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg 6421 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt 6481 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac 6541 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg 6601 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg 6661 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat 6721 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt 6781 tacggttcct ggccttttgc tggccttttg ctcacatgt //