LOCUS       Exported                5116 bp ds-DNA     circular SYN 01-JUL-2016
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    9073_EGFPC1_U6-utarget7-spacer_ultimate target no7
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5116)
  AUTHORS   Talas Andras
  TITLE     Direct Submission
  JOURNAL   Exported Friday, July 1, 2016 from SnapGene 3.1.4
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5116
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     enhancer        61..364
                     /note="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /note="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             613..1329
                     /codon_start=1
                     /product="enhanced GFP"
                     /note="EGFP"
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     misc_feature    1330..1395
                     /note="MCS"
                     /note="multiple cloning site of fluorescent protein 
                     plasmids"
     misc_feature    1404..1414
                     /note="stop codons"
                     /note="stop codons in all three reading frames"
     polyA_signal    1519..1640
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1647..2102)
                     /direction=LEFT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        2129..2233
                     /gene="bla"
                     /note="AmpR promoter"
     promoter        2235..2592
                     /note="SV40 promoter"
                     /note="SV40 enhancer and early promoter"
     CDS             2627..3421
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /note="NeoR/KanR"
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin)"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     misc_feature    3653..3700
                     /note="HSV TK poly(A) signal"
                     /note="herpesvirus thymidine kinase polyadenylation signal"
     promoter        3754..3986
                     /label=hU6_promoter
                     /note="hU6_promoter"
     misc_feature    3994..4013
                     /note="spacer"
     misc_RNA        4014..4089
                     /note="gRNA scaffold"
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     rep_origin      4380..4968
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    5033..5055
                     /note="ultimate target no 7"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attctgcagt cgacggtacc
     1381 gcgggcccgg gatccaccgg atctagataa ctgatcataa tcagccatac cacatttgta
     1441 gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg
     1501 aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     1561 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     1621 aaactcatca atgtatctta acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc
     1681 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     1741 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     1801 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     1861 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     1921 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     1981 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     2041 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     2101 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     2161 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     2221 aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt
     2281 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     2341 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     2401 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     2461 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     2521 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     2581 ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     2641 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     2701 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     2761 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc
     2821 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     2881 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     2941 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     3001 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     3061 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     3121 cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt
     3181 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     3241 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     3301 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     3361 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     3421 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     3481 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     3541 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg
     3601 aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata
     3661 aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc
     3721 cagggctggc actctgtcga taccccacgc ctatttccca tgattccttc atatttgcat
     3781 atacgataca aggctgttag agagataatt ggaattaatt tgactgtaaa cacaaagata
     3841 ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa
     3901 ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg
     3961 gctttatata tcttgtggaa aggacgaaac accgatcaat ttcgccgcaa cgcgttttag
     4021 agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa agtggcaccg
     4081 agtcggtgct tttttccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt
     4141 tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg
     4201 gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat
     4261 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4321 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4381 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4441 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4501 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4561 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4621 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4681 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4741 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4801 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4861 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4921 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4981 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgct ccgatcaatt
     5041 tcgccgcaac gccggtagca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     5101 tattaccgcc atgcat
//
