LOCUS       pEV712                  3749 bp    DNA     circular SYN 18-JAN-2021
DEFINITION  cHS4-2xgRNA1 BS-miniTK-EYFP-SV40.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3749)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 25, 2025 from SnapGene 8.1.1
            https://www.snapgene.com
COMMENT     Imported From
            Path: C:\Users\UPALTO~1\AppData\Local\Temp\Rar$DRa0.846
            Filename: pL1F-1-s1-s2_RFP.ape
FEATURES             Location/Qualifiers
     source          1..3749
                     /lab_host="."
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     source          2682..2689
                     /mol_type="other DNA"
                     /note="color: #ffffff"
                     /organism="synthetic DNA construct"
     misc_feature    1..45
                     /label=Seq2
                     /note="/ApEinfo_fwdcolor=#8000ff"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1..45
                     /label=seq2
                     /note="/ApEinfo_fwdcolor=#6666ff"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(40..45)
                     /label=BBb scar(4)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: LEFT"
     protein_bind    54..71
                     /label=I-SceI
                     /note="/created by=User"
                     /note="color: #31849b; direction: RIGHT"
     misc_feature    92..201
                     /label=RK2 trfa
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    206..211
                     /label=BBb scar
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     rep_origin      complement(229..911)
                     /direction=LEFT
                     /label=ColE1 origin
                     /note="/ApEinfo_fwdcolor=gray50"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_revcolor=gray50"
                     /note="color: #ffff00"
     misc_feature    827..832
                     /label=BBb scar(1)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(838..843)
                     /label=BBb scar(5)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    924..929
                     /label=BBb scar(2)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(936..941)
                     /label=BBb scar(6)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    1009..1866
                     /label=AmpR
                     /note="/ApEinfo_fwdcolor=#ff80ff"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1704..1709
                     /label=BBb scar(3)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(1721..1726)
                     /label=BBb scar(7)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: LEFT"
     protein_bind    complement(1992..2009)
                     /label=I-SceI
                     /note="/created by=User"
                     /note="color: #31849b; direction: LEFT"
     misc_feature    2013..2057
                     /label=seq1
                     /note="/ApEinfo_fwdcolor=#8000ff"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: RIGHT"
     protein_bind    2058..2061
                     /label=GGAG
                     /note="/created by=elviravitu"
                     /note="/modified by=elviravitu"
                     /note="color: #31849b"
     misc_feature    2062..2591
                     /label=cHS4 Insulator x2
                     /note="Geneious type: chromatin insulator"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(2062..2591)
                     /label=Digestion of pZDonor-L_ISceI-Seq1-FseI-SeqX - F...
                     /note="Geneious type: Concatenated sequence"
                     /note="/NCBI Join Type=order"
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    2062..2591
                     /label=Digestion of MAV1213-Hef1a-mKate2 - Fragment 1 ...
                     /note="Geneious type: Concatenated sequence"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2062..2591
                     /label=Digestion of MAV1212_Hef1a-mKate2 (reduced AarI...
                     /note="Geneious type: Concatenated sequence"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2323..2326
                     /label=AgeI
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2341..2591
                     /label=cHS4 Core
                     /note="color: #a6acb3; direction: RIGHT"
     protein_bind    2612..2615
                     /label=TACT
                     /note="/created by=User"
                     /note="color: #31849b"
     rRNA            2616..2635
                     /label=gRNA1 BS
                     /note="/created by=User"
                     /note="/modified by=elviravitu"
                     /note="color: #00ff00; direction: RIGHT"
     misc_feature    2636..2638
                     /label=PAM
                     /note="Geneious type: motif"
                     /note="/created by=User"
                     /note="/modified by=User"
                     /note="color: #a6acb3"
     rRNA            2657..2676
                     /label=gRNA1 BS
                     /note="/created by=User"
                     /note="/modified by=elviravitu"
                     /note="color: #00ff00; direction: RIGHT"
     misc_feature    2677..2679
                     /label=PAM
                     /note="Geneious type: motif"
                     /note="/created by=User"
                     /note="/modified by=User"
                     /note="color: #a6acb3"
     promoter        2690..2752
                     /label=minTK
                     /note="/created by=elviravitu"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    2753..2756
                     /label=AATG
                     /note="/created by=User"
                     /note="color: #31849b"
     unsure          2757..2795
                     /label=5' UTR
                     /note="/created by=elviravitu"
                     /note="color: #a6acb3"
     protein_bind    2796..2799
                     /label=AGGT
                     /note="/created by=User"
                     /note="color: #31849b"
     misc_feature    2800..2805
                     /label=Kozak
                     /note="Geneious type: motif"
                     /note="color: #a6acb3; direction: RIGHT"
     gene            2806..3525
                     /label=eYFP
                     /note="/modified by=elviravitu"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(3305..3310)
                     /label=BBb scar(2)
                     /note="/ApEinfo_fwdcolor=cyan"
                     /note="/ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {}
                     0} width 5 offset 0"
                     /note="/ApEinfo_label=BBb scar"
                     /note="/ApEinfo_revcolor=green"
                     /note="color: #a6acb3; direction: LEFT"
     protein_bind    3526..3529
                     /label=GCTT
                     /note="/created by=User"
                     /note="color: #31849b"
     protein_bind    3550..3553
                     /label=CAAC
                     /note="/created by=User"
                     /note="color: #31849b"
     polyA_signal    3554..3745
                     /label=SV40 polyA
                     /note="/created by=User"
                     /note="/modified by=elviravitu"
                     /note="color: #a6acb3; direction: RIGHT"
     protein_bind    3746..3749
                     /label=CGCT
                     /note="/created by=User"
                     /note="color: #31849b"
ORIGIN
        1 ggtgcgtttt tatgcttgta gtattgtata atgtttttaa gatccttaat taatagggat
       61 aacagggtaa tatcttgccc tgcacgaata ccagcgaccc cttgcccaaa tacttgccgt
      121 gggcctcggc ctgagagcca aaacacttga tgcggaagaa gtcggtgcgc tcctgcttgt
      181 cgccggcatc gttgcgccac atctaggatc tgccaggaac cgtaaaaagg ccgcgttgct
      241 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
      301 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
      361 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
      421 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
      481 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
      541 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
      601 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
      661 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
      721 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
      781 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
      841 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
      901 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
      961 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     1021 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     1081 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     1141 accgcgagaa ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     1201 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     1261 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     1321 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     1381 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     1441 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     1501 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     1561 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     1621 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     1681 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     1741 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     1801 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     1861 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     1921 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacgaattg
     1981 gccagcgctg cattaccctg ttatccctaa ccggtttacc gagctcttat tggttttcaa
     2041 acttcattga ctgtgccgga ggcgcctggg agctcacggg gacagccccc ccccaaagcc
     2101 cccagggatg taattacgtc cctcccccgc tagggggcag cagcgagccg cccggggctc
     2161 cgctccggtc cggcgctccc cccgcatccc cgagccggca gcgtgcgggg acagcccggg
     2221 cacggggaag gtggcacggg atcgctttcc tctgaacgct tctcgctgct ctttgagcct
     2281 gcagacacct ggggggatac ggggaaaaag ctaggctgaa agaggttctt agaatgacac
     2341 gcgcctggga gctcacgggg acagcccccc cccaaagccc ccagggatgt aattacgtcc
     2401 ctcccccgct agggggcagc agcgagccgc ccggggctcc gctccggtcc ggcgctcccc
     2461 ccgcatcccc gagccggcag cgtgcgggga cagcccgggc acggggaagg tggcacggga
     2521 tcgctttcct ctgaacgctt ctcgctgctc tttgagcctg cagacacctg gggggatacg
     2581 gggaaaaagc tggatcataa ctcgacgtgc atactgtccc ctccacccca cagtggggca
     2641 tagcgccggc cgcgccgtcc cctccacccc acagtggggc accatatgct tcgcatatta
     2701 aggtgacgcg tgtggcctcg aacaccgagc gaccctgcag cgacccgctt aaaatgctag
     2761 tctactacca gagctcatcg ctagcgctac cggtcaggtg ccaccatggt gagcaagggc
     2821 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     2881 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     2941 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccttc
     3001 ggctacggcc tgcagtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc
     3061 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     3121 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     3181 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     3241 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     3301 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     3361 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagctaccag
     3421 tccaagctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     3481 accgccgccg ggatcactct cggcatggac gagctgtaca agtgagcttc cactggattg
     3541 tacaattacc aaccagacat gataagatac attgatgagt ttggacaaac cacaactaga
     3601 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     3661 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
     3721 cagggggagg tgtgggaggt tttttcgct
//
