LOCUS Exported 7245 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Express Ran Q69L mutant in mammalian cells (CMV promoter). Also suitable for in vitro transcription (T7 promoter) for injecting mRNA into oocytes.. ACCESSION . VERSION . KEYWORDS pcDNA-Ran-Q69L-mRFP1-polyA SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7245) AUTHORS Inoue A, Zhang Y TITLE Nucleosome assembly is required for nuclear pore complex assembly in mouse zygotes. JOURNAL Nat Struct Mol Biol. 2014 Jul;21(7):609-16. doi: 10.1038/nsmb.2839. Epub 2014 Jun 8. PUBMED 24908396 REFERENCE 2 (bases 1 to 7245) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7245 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 171..550 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 551..754 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 705..725 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 799..818 /label=T7 /note="T7 promoter, forward primer" promoter 799..817 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 1498..2175 /codon_start=1 /product="monomeric derivative of DsRed (Campbell et al., 2002)" /label=mRFP1 /note="mammalian codon-optimized" /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS TGA" primer_bind complement(1628..1646) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 1896..1915 /label=mCherry-F /note="mCherry, forward primer" primer_bind 2090..2113 /label=DsRed1-C /note="DsRed1, forward primer" CDS 2685..2714 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" CDS 2730..2747 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(2770..2787) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 2776..3000 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 3046..3474 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3133..3152) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3343..3364 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(3483..3503) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3488..3817 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3668..3803 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3730..3749 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3884..4678 /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(3938..3957) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4548..4567 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 4854..4975 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4891..4910) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4945..4964 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(5024..5040) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5024..5040) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5037..5059) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5048..5064 /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(5072..5102) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5117..5138 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5255..5272) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5426..6014) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5506..5525) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6185..7045) /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 6808..6827 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7046..7150) /gene="bla" /label=AmpR promoter primer_bind complement(7225..7244) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag 61 caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag 121 ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt gacattgatt 181 attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga 241 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg 301 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg 361 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca 421 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc 481 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc 541 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc 601 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa 661 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag 721 gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag aacccactgc 781 ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg gctagttaag 841 cttatggccg cccagggaga gccgcaggtc cagttcaagc tcgtcctggt gggcgacggc 901 ggcaccggga agacaacctt cgtgaagcgc cacttgacgg gcgagtttga gaagaagtat 961 gtagccaccc tgggcgtgga ggtgcacccg ctcgtcttcc ataccaacag aggacccatc 1021 aagttcaacg tgtgggacac ggccggcctg gagaagttcg ggggcctgcg cgatggctac 1081 tacatccaag cccagtgtgc cattataatg tttgatgtaa cctcaagagt tacttacaag 1141 aatgtaccta actggcatag agatctggta cgagtgtgtg aaaacatccc cattgtattg 1201 tgtggcaaca aagtggatat taaagacagg aaagtgaagg caaaatctat tgtcttccac 1261 cggaagaaga atcttcagta ctatgacatt tctgccaaaa gtaactacaa ctttgaaaag 1321 cctttcctct ggcttgccag aaagctcatt ggagatccta acttggagtt tgttgccatg 1381 cctgctcttg ccccacctga ggtggtcatg gacccagctt tggcagcaca gtacgagcat 1441 gatttagagg ttgctcagac gactgctctc ccagatgagg atgatgacct ggatccaatg 1501 gcctcctccg aggacgtcat caaggagttc atgcgcttca aggtgcgcat ggagggctcc 1561 gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc 1621 cagaccgcca agctgaaggt gaccaagggc ggccccctgc ccttcgcctg ggacatcctg 1681 tcccctcagt tccagtacgg ctccaaggcc tacgtgaagc accccgccga catccccgac 1741 tacttgaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac 1801 ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt catctacaag 1861 gtgaagctgc gcggcaccaa cttcccctcc gacggccccg taatgcagaa gaagaccatg 1921 ggctgggagg cctccaccga gcggatgtac cccgaggacg gcgccctgaa gggcgagatc 1981 aagatgaggc tgaagctgaa ggacggcggc cactacgacg ccgaggtcaa gaccacctac 2041 atggccaaga agcccgtgca gctgcccggc gcctacaaga ccgacatcaa gctggacatc 2101 acctcccaca acgaggacta caccatcgtg gaacagtacg agcgcgccga gggccgccac 2161 tccaccggcg cctaagcggc cgctcgacta tgggcaccaa agaacctgta aacgttatct 2221 ttttaaattg aatgtgcaca aataaaagtt tggaaaagaa aaaaaaaaaa aaaaaaaaaa 2281 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaamaaa 2341 aaaaaaamaa aaamaaaaam aaaaaaaaan aaaaaaaaaa aaaaaaaaaa aamaaaaaaa 2401 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aannnnaaaa aaaaaaaaaa aaaaaaaaaa 2461 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa 2521 aaaaaaaaaa naaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa 2581 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaat 2641 acgcatcata cgctgtggcg tacctcgagt ctagagggcc cttcgaacaa aaactcatct 2701 cagaagagga tctgaatatg cataccggtc atcatcacca tcaccattga gtttaaaccc 2761 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg 2821 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa 2881 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca 2941 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg 3001 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg 3061 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg 3121 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc 3181 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc 3241 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga 3301 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa 3361 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga 3421 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct 3481 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat 3541 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc 3601 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac 3661 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact 3721 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta 3781 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc 3841 cattttcgga tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg 3901 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca 3961 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt 4021 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg 4081 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga 4141 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca 4201 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct 4261 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac 4321 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc 4381 gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt 4441 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt 4501 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg 4561 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat 4621 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc 4681 gggactctgg ggttcgcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt 4741 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg 4801 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt 4861 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag 4921 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg 4981 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg 5041 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta 5101 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg 5161 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga 5221 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg 5281 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag 5341 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc 5401 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca 5461 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt 5521 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc 5581 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc 5641 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc 5701 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact 5761 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg 5821 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta 5881 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca 5941 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa 6001 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg 6061 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc 6121 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg 6181 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat 6241 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg 6301 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa 6361 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca 6421 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc 6481 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt 6541 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa 6601 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat 6661 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct 6721 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga 6781 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag 6841 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga 6901 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca 6961 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg 7021 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc 7081 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag 7141 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc 7201 cgatccccta tggtgcactc tcagtacaat ctgctctgat gccgc //