LOCUS Exported 5863 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expresses myc-M87E112K spastin isoform in mammalian cells. ACCESSION . VERSION . KEYWORDS pCMV-Tag 3B/myc-M87E112K SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5863) AUTHORS Solowska JM, Garbern JY, Baas PW TITLE Evaluation of loss of function as an explanation for SPG4-based hereditary spastic paraplegia. JOURNAL Hum Mol Genet. 2010 Jul 15;19(14):2767-79. doi: 10.1093/hmg/ddq177. Epub 2010 Apr 29. PUBMED 20430936 REFERENCE 2 (bases 1 to 5863) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5863 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..304 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 305..508 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 459..479 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" promoter 554..572 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" regulatory 604..613 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 613..642 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" primer_bind complement(2298..2317) /label=T7 /note="T7 promoter, forward primer" promoter complement(2299..2317) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" polyA_signal 2591..2712 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2628..2647) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2682..2701 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2719..3174) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2856..2877) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3068..3087 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3201..3305 /gene="bla" /label=AmpR promoter promoter 3307..3664 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3330..3350) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3515..3650 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3577..3596 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3699..4493 /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 GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3753..3772) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4363..4382 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4681..4700) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4725..4772 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 5101..5689 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5590..5609 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgatt 541 acgccaagct cgaaattaac cctcactaaa gggaacaaaa gctggagctc caccgcggtg 601 gcggccgcca tggagcagaa actcatctct gaagaggatc tgagcccggg cggatccgcc 661 gccaagagga gctccggggc cgcgccagca cctgcctcgg cctcggcccc ggcgccggtg 721 ccgggcggca aggccgagcg cgtccgagtc ttccacaaac aggccttcga gtacatctcc 781 attgccctgc gcatcgatga ggatgagaaa gcaggacaga aggagcaagc tgtggaatgg 841 tataagaaag gtattgaaga actggaaaaa ggaatagctg ttatagttac aggacaaggt 901 gaacagtgtg aaagagctag acgccttcaa gctaaaatga tgactaattt ggttatggcc 961 aaggaccgct tacaacttct agagaagatg caaccagttt tgccattttc caagtcacaa 1021 acggacgtct ataatgacag tactaacttg gcatgccgca atggacatct ccagtcagaa 1081 agtggagctg ttccaaaaag aaaagacccc ttaacacaca ctagtaattc actgcctcgt 1141 tcaaaaacag ttatgaaaac tggatctgca ggcctttcag gccaccatag agcacctagt 1201 tacagtggtt tatccatggt ttctggagtg aaacagggat ctggtcctgc tcctaccact 1261 cataagggta ctccgaaaac aaataggaca aataaacctt ctacccctac aactgctact 1321 cgtaagaaaa aagacttgaa gaattttagg aatgtggaca gcaaccttgc taaccttata 1381 atgaatgaaa ttgtggacaa tggaacagct gttaaatttg atgatatagc tggtcaagac 1441 ttggcaaaac aagcattgca agaaattgtt attcttcctt ctctgaggcc tgagttgttc 1501 acagggctta gagctcctgc cagagggctg ttactctttg gtccacctgg gaatgggaag 1561 acaatgctgg ctaaagcagt agctgcagaa tcgaatgcaa ccttctttaa tataagtgct 1621 gcaagtttaa cttcaaaata cgtgggagaa ggagagaaat tggtgagggc tctttttgct 1681 gtggctcgag aacttcaacc ttctataatt tttatagatg aagttgatag ccttttgtgt 1741 gaaagaagag aaggggagca cgatgctagt agacgcctaa aaactgaatt tctaatagaa 1801 tttgatggtg tacagtctgc tggagatgac agagtacttg taatgggtgc aactaatagg 1861 ccacaagagc ttgatgaggc tgttctcagg cgtttcatca aacgggtata tgtgtcttta 1921 ccaaatgagg agacaagact acttttgctt aaaaatctgt tatgtaaaca aggaagtcca 1981 ttgacccaaa aagaactagc acaacttgct agaatgactg atggatactc aggaagtgac 2041 ctaacagctt tggcaaaaga tgcagcactg ggtcctatcc gagaactaaa accagaacag 2101 gtgaagaata tgtctgccag tgagatgaga aatattcgat tatctgactt cactgaatcc 2161 ttgaaaaaaa taaaacgcag cgtcagccct caaactttag aagcgtacat acgttggaac 2221 aaggactttg gagataccac tgtttaaggg cccggtacct taattaatta aggtaccagg 2281 taagtgtacc caattcgccc tatagtgagt cgtattacaa ttcactcgat cgcccttccc 2341 aacagttgcg cagcctgaat ggcgaatgga gatccaattt ttaagtgtat aatgtgttaa 2401 actactgatt ctaattgttt gtgtatttta gattcacagt cccaaggctc atttcaggcc 2461 cctcagtcct cacagtctgt tcatgatcat aatcagccat accacatttg tagaggtttt 2521 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat 2581 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac 2641 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat 2701 caatgtatct taacgcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt 2761 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat 2821 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat 2881 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac 2941 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc 3001 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga 3061 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca 3121 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg 3181 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa 3241 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga 3301 agaatcctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc 3361 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag 3421 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta 3481 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc 3541 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc 3601 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg 3661 caaagatcga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc 3721 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga 3781 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt 3841 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat 3901 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg 3961 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg 4021 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc 4081 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga 4141 tggaagccgg tcttgtcgat caggatgatc tggacgaaga acatcagggg ctcgcgccag 4201 ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc 4261 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg 4321 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata 4381 ttgctgaaga acttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg 4441 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac 4501 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc 4561 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat 4621 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac 4681 tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca 4741 gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg 4801 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt 4861 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc 4921 ggcaggccct gccatagcct caggttactc atatatactt tagattgatt taaaacttca 4981 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc 5041 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc 5101 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc 5161 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt 5221 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt 5281 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc 5341 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa 5401 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac 5461 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg 5521 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga 5581 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact 5641 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa 5701 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc 5761 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat 5821 caattacggg gtcattagtt catagcccat atatggagtt ccg //