LOCUS Exported 5701 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION destination vector for the Golden Gate TALEN kit, directs expression of TALENs from a truncated CAGs promoter. ACCESSION . VERSION . KEYWORDS pC-GoldyTALEN SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5701) AUTHORS Carlson DF, Tan W, Lillico SG, Stverakova D, Proudfoot C, Christian M, Voytas DF, Long CR, Whitelaw CB, Fahrenkrug SC TITLE Efficient TALEN-mediated gene knockout in livestock. JOURNAL Proc Natl Acad Sci U S A. 2012 Oct 1. PUBMED 23027955 REFERENCE 2 (bases 1 to 5701) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5701 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 104..407 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 409..684 /label=chicken beta-actin promoter primer_bind 940..959 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" CDS 1062..1082 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 1083..1109 /codon_start=1 /product="epitope tag from the baculovirus Autographa californica GP64 envelope fusion protein" /label=AcV5 tag /translation="SWKDASGWS" CDS complement(1413..1820) /codon_start=1 /gene="lacZ fragment" /product="LacZ-alpha fragment of beta-galactosidase" /label=lacZ-alpha /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLQGG TRQCIARCMHCLHCGHAATFCNLHKLACVQL" primer_bind 1670..1692 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 1684..1701 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 1685..1701 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 1711..1730 /label=T7 /note="T7 promoter, forward primer" promoter 1711..1729 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1777..1797) /label=T3 /note="T3 promoter, forward primer" promoter complement(1777..1795) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(1816..1832) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1816..1832) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1829..1851) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1840..1856 /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(1864..1894) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1909..1930 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." CDS 2119..2706 /codon_start=1 /product="nonspecific DNA cleavage domain of the FokI endonuclease (Li et al., 1992)" /label=FokI cleavage domain /note="must dimerize to be catalytically active" /translation="QLVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKV MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMQR YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF" polyA_signal 2817..2898 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" polyA_signal 3022..3103 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" rep_origin complement(3271..3726) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3408..3429) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3620..3639 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3753..3857 /gene="bla" /label=AmpR promoter CDS 3858..4718 /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 complement(4076..4095) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4889..5477 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5378..5397 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5631..5648 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 tcattaatgc agctgagatc ttaagggatt actagtttaa acctagttat taatagtaat 61 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg 121 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt 181 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac 241 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg 301 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact 361 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtc gaggtgagcc 421 ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt ttgtatttat 481 ttatttttta attattttgt gcagcgatgg gggcgggggg ggggggggcg cgcgccaggc 541 ggggcggggc ggggcgaggg gcggggcggg gcgaggcgga gaggtgcggc ggcagccaat 601 cagagcggcg cgctccgaaa gtttcctttt atggcgaggc ggcggcggcg gcggccctat 661 aaaaagcgaa gcgcgcggcg ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc 721 tccgccgccg cctcgcgccg cccgccccgg ctctgactga ccgcgttact cccacaggtg 781 agcgggcggg acggcccttc tcctccgggc tgtaattagc gcttggttta atgacggctt 841 gtttcttttc tgtggctgcg tgaaagcctt gaggggctcc gggagctaga gcctctgcta 901 accatgttca tgccttcttc tttttcctac agctcctggg caacgtgctg gttattgtgc 961 tgtctcatca ttttggcaaa gaattttaag ctaggatcta actcgagaaa gatattgtat 1021 atatcgtaac aataggaggt tcaacaatgg cttcctcccc tccaaagaaa aagagaaagg 1081 ttagttggaa ggacgcaagt ggttggtcta gagtggatct acgcacgctc ggctacagtc 1141 agcagcagca agagaagatc aaaccgaagg tgcgttcgac agtggcgcag caccacgagg 1201 cactggtggg ccatgggttt acacacgcgc acatcgttgc gctcagccaa cacccggcag 1261 cgttagggac cgtcgctgtc acgtatcagc acataatcac ggcgttgcca gaggcgacac 1321 acgaagacat cgttggcgtc ggcaaacagt ggtccggcgc acgcgccctg gaggccttgc 1381 tcacggatgc gggggagttg agaggtccgc cgttacagtt ggacacaggc caacttgtga 1441 agattgcaaa acgtggcggc gtgaccgcaa tggaggcagt gcatgcatcg cgcaatgcac 1501 tgacgggtgc ccccctggag acgggcgccg ctacagggcg cgtcccattc gccattcagg 1561 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg 1621 aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga 1681 cgttgtaaaa cgacggccag tgagcgcgcg taatacgact cactataggg cgaattgggt 1741 accgggcccc ccctcgaggt cctccagctt ttgttccctt tagtgagggt taattgcgcg 1801 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc 1861 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta 1921 actcacatta attgcgttgc gctcactgcc cgctttccac cggtcgtctc caacgaccac 1981 ctcgtcgcct tggcctgcct cggcggacgt cctgccatgg atgcagtgaa aaagggattg 2041 ccgcacgcgc cggaattgat cagaagagtc aatcgccgta ttggcgaacg cacgtcccat 2101 cgcgttgcct ctagatccca gctagtgaaa tctgaattgg aagagaagaa atctgaactt 2161 agacataaat tgaaatatgt gccacatgaa tatattgaat tgattgaaat cgcaagaaat 2221 tcaactcagg atagaatcct tgaaatgaag gtgatggagt tctttatgaa ggtttatggt 2281 tatcgtggta aacatttggg tggatcaagg aaaccagacg gagcaattta tactgtcgga 2341 tctcctattg attacggtgt gatcgttgat actaaggcat attcaggagg ttataatctt 2401 ccaattggtc aagcagatga aatgcaaaga tatgtcgaag agaatcaaac aagaaacaag 2461 catatcaacc ctaatgaatg gtggaaagtc tatccatctt cagtaacaga atttaagttc 2521 ttgtttgtga gtggtcattt caaaggaaac tacaaagctc agcttacaag attgaatcat 2581 atcactaatt gtaatggagc tgttcttagt gtagaagagc ttttgattgg tggagaaatg 2641 attaaagctg gtacattgac acttgaggaa gtgagaagga aatttaataa cggtgagata 2701 aacttttaat aggagctcgc ccggggatct aattcaatta gagactaatt caattagagc 2761 taattcaatt aggatccaga attcattgat gagtttggac aaaccacaac tagaatgcag 2821 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata 2881 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg 2941 gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtca ttgatgagtt 3001 tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc 3061 tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat 3121 tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct 3181 ctacaaatgt ggtcattcag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc 3241 caacagttgc gcagcctgaa tggcgaatgg aaattgtaag cgttaatatt ttgttaaaat 3301 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa 3361 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca 3421 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg 3481 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta 3541 aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg 3601 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa 3661 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg 3721 gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct 3781 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat 3841 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg 3901 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg 3961 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc 4021 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat 4081 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact 4141 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca 4201 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact 4261 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg 4321 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg 4381 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg 4441 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg 4501 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag 4561 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc 4621 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga 4681 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat 4741 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc 4801 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag 4861 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct 4921 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac 4981 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc 5041 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg 5101 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt 5161 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt 5221 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc 5281 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca 5341 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata 5401 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg 5461 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct 5521 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta 5581 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag 5641 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga 5701 t //