LOCUS       pFC482                  4564 bp    DNA     circular SYN 10-MAR-2020
DEFINITION  cHS4-hEF1a-5'UTR-NLS-CasE-3'UTR-terminator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4564)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 25, 2025 from SnapGene 8.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4564
                     /lab_host="Cloning vector pUC19"
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     source          2945..3569
                     /mol_type="other DNA"
                     /note="color: #ffffff"
                     /organism="Cloning vector pUC19"
     source          2952..2975
                     /mol_type="other DNA"
                     /note="color: #ffffff"
                     /organism="synthetic DNA construct"
     misc_feature    13..18
                     /label=BBb scar
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(25..30)
                     /label=BBb scar
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    complement(95..955)
                     /label=amp marker
                     /note="Geneious type: marker"
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    98..955
                     /label=AmpR
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    793..798
                     /label=BBb scar
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(810..815)
                     /label=BBb scar
                     /note="color: #a6acb3; direction: LEFT"
     promoter        complement(997..1025)
                     /label=amp prom
                     /note="color: #ffffff; direction: LEFT"
     misc_feature    1102..1146
                     /label=seq1
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1147..1724
                     /label=pBjh_3260 (concatenated sequence 2)
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1147..1150
                     /label=pL0 I
                     /note="Geneious type: gg_overhang"
                     /note="/created_by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1147..1150
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1147..1150
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3"
     misc_feature    1147..1150
                     /label=ligation
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1151..1720
                     /label=2x cHS4 Insulator
                     /note="Geneious type: Insulator"
                     /note="/created_by=User"
                     /note="/modified_by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     protein_bind    1229..1233
                     /label=CTCF Bind (Short)
                     /note="/created_by=User"
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    1508..1512
                     /label=CTCF Bind (Short)
                     /note="/created_by=User"
                     /note="color: #31849b; direction: RIGHT"
     misc_feature    1721..2918
                     /label=L0.P.2_hEF1a (concatenated sequence 3)
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1721..1724
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1721..1724
                     /label=pL0 I-P
                     /note="Geneious type: gg_overhang"
                     /note="/created_by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    1721..1724
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3"
     misc_feature    1721..1724
                     /label=pL0 I-P
                     /note="Geneious type: gg_overhang"
                     /note="color: #a6acb3; direction: RIGHT"
     promoter        1729..1961
                     /label=hEF1a Promoter
                     /note="color: #ffffff; direction: RIGHT"
     enhancer        1758..2019
                     /label=CpG Island
                     /note="color: #a6acb3; direction: RIGHT"
     promoter        1893..1898
                     /label=BRE-like
                     /note="color: #ffffff; direction: RIGHT"
     promoter        1899..1905
                     /label=TATA Box
                     /note="color: #ffffff; direction: RIGHT"
     misc_RNA        1929
                     /label=+1 Site
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(1943..1961)
                     /label=RJ_00293Ra
                     /note="color: #a6acb3; direction: LEFT"
     intron          1959..1967
                     /label=U2 Splice Donor
                     /note="color: #------; direction: RIGHT"
     primer_bind     1960..1978
                     /label=RJ_00296Fa
                     /note="color: black; sequence: AGGTAAGTGCCGTGTGTGG"
     misc_RNA        1961..1962
                     /label=Cut
                     /note="color: #a6acb3; direction: RIGHT"
     intron          1962..2900
                     /label=hEF1a First Intron
                     /note="color: #------; direction: RIGHT"
     enhancer        2125..2214
                     /label=CpG Island
                     /note="color: #a6acb3; direction: RIGHT"
     enhancer        2317..2717
                     /label=CpG Island
                     /note="color: #a6acb3; direction: RIGHT"
     protein_bind    2317..2331
                     /label=Gal4 Close Match (15)
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    2332..2350
                     /label=Gal4 Close Match (19)
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    2333..2338
                     /label=SP1 Binding Site
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    2336..2350
                     /label=Gal4 Close Match (15)
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    complement(2475..2480)
                     /label=SP1 Binding Site
                     /note="color: #31849b; direction: LEFT"
     protein_bind    2482..2487
                     /label=SP1 Binding Site
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    2485..2500
                     /label=Gal4 Close Match (16)
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    2590..2595
                     /label=SP1 Binding Site
                     /note="color: #31849b; direction: RIGHT"
     misc_RNA        2595..2596
                     /label=Putative Cut
                     /note="color: #a6acb3; direction: RIGHT"
     protein_bind    2597..2603
                     /label=AP-1 Binding Site
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    2659..2675
                     /label=Gal4 Close Match
                     /note="color: #31849b; direction: RIGHT"
     protein_bind    complement(2750..2759)
                     /label=NFkB Binding Site
                     /note="color: #31849b; direction: LEFT"
     intron          2865..2869
                     /label=U2 Branch Point-like
                     /note="color: #------; direction: RIGHT"
     intron          2882..2897
                     /label=Poly-Y Tract
                     /note="color: #------; direction: RIGHT"
     misc_feature    complement(2889..2914)
                     /label=7.21
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    complement(2889..2909)
                     /label=RJ_00296Ra
                     /note="color: #a6acb3; direction: LEFT"
     intron          2898..2901
                     /label=U2 Splice Acceptor
                     /note="color: #------; direction: RIGHT"
     misc_RNA        2900..2901
                     /label=Cut
                     /note="color: #a6acb3; direction: RIGHT"
     5'UTR           2901..2914
                     /label=hEF1a 5'UTR
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2915..2942
                     /label=L0.5.2_Inert (concatenated sequence 4)
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2915..2918
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2915..2918
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3"
     misc_feature    2915..2918
                     /label=pL0 P-5 v2
                     /note="Geneious type: gg_overhang"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2915..2918
                     /label=ligation
                     /note="color: #a6acb3; direction: RIGHT"
     5'UTR           2919..2938
                     /label=Inert
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2939..3576
                     /label=pFC477 (concatenated sequence 5)
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     primer_bind     2939..2999
                     /label=PRFC277
                     /note="color: black; sequence: 
                     AAAGAAGACTTAGGTGCCACCATGAGCCCCAAGAAGAAGAGAAAGGTGTACCTCAGTAA
                     GATCATCATCGCC"
     misc_feature    2939..2942
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3"
     misc_feature    2939..2942
                     /label=ligation
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2943..3572
                     /label=Digestion of pKL451 extraction - Fragment 2 ext...
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2943..2944
                     /label=Kozak
                     /note="/created by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2945..2948
                     /label=BsaI OH
                     /note="/created by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2949..3569
                     /label=Cse3
                     /note="Geneious type: ORF"
                     /note="/created by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     CDS             2955..2975
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /note="Geneious type: motif"
                     /note="/modified by=User"
                     /note="color: #993366"
                     /translation="PKKKRKV"
     primer_bind     complement(3077..3093)
                     /label=WX-139R
                     /note="color: black; sequence: CGTGGAACAGGAAGTCT"
     primer_bind     3437..3454
                     /label=WX-138F
                     /note="color: black; sequence: CGGCAAAATTCAGACTGT"
     primer_bind     complement(3547..3569)
                     /label=Primer 1
                     /note="color: black; sequence: CAGGGGTGCCAGGCTCAACAATC"
     primer_bind     complement(3554..3576)
                     /label=PRFC278
                     /note="color: black; sequence: 
                     AAAGAAGACTTAAGCTCACAGGGGTGCCAGGCTC"
     misc_feature    3573..3600
                     /label=L0.3_Inert (concatenated sequence 6)
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3573..3576
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3"
     misc_feature    3573..3576
                     /label=ligation
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3577..3596
                     /label=Inert
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3597..3653
                     /label=pBjh_4560 (concatenated sequence 7)
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3597..3600
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3"
     misc_feature    3597..3600
                     /label=ligation
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3597..3600
                     /label=pL0 3-T
                     /note="Geneious type: gg_overhang"
                     /note="/created_by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     terminator      3601..3649
                     /label=Synth Terminator (Short)
                     /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="/modified_by=User"
                     /note="color: #ffffff; direction: RIGHT"
     polyA_signal    3601..3606
                     /label=Poly-A Signal
                     /note="/created_by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     polyA_signal    complement(3612..3617)
                     /label=Poly-A Signal
                     /note="/created_by=User"
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    3650..5714
                     /label=L1.Seq1-Seq2_Backbone (concatenated sequence 1)
                     /note="Geneious type: Concatenated sequence"
                     /note="/Source"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3650..3653
                     /label=Ligation
                     /note="Geneious type: ligation"
                     /note="color: #a6acb3"
     misc_feature    3650..3653
                     /label=pL0 T
                     /note="Geneious type: gg_overhang"
                     /note="/created_by=User"
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3650..3653
                     /label=ligation
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3654..3698
                     /label=seq2
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(3693..3698)
                     /label=BBb scar
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    3745..3854
                     /label=RK2 trfa
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3859..3864
                     /label=BBb scar
                     /note="color: #a6acb3; direction: RIGHT"
     rep_origin      complement(3882..4564)
                     /direction=LEFT
                     /label=ColE1 origin
                     /note="color: #ffff00"
     misc_feature    complement(3885..4504)
                     /label=pBR322 origin
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    4480..4485
                     /label=BBb scar
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    complement(4491..4496)
                     /label=BBb scar
                     /note="color: #a6acb3; direction: LEFT"
ORIGIN
        1 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
       61 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
      121 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
      181 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagaac
      241 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
      301 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
      361 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
      421 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
      481 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
      541 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
      601 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
      661 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
      721 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
      781 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
      841 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
      901 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
      961 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     1021 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacgaattgg ccagcgctgc
     1081 attaccctgt tatccctaac cggtttaccg agctcttatt ggttttcaaa cttcattgac
     1141 tgtgccggag ttctgcaatt catgtgatct gcgcctggga gctcacgggg acagcccccc
     1201 cccaaagccc ccagggatgt aattacgtcc ctcccccgct agggggcagc agcgagccgc
     1261 ccggggctcc gctccggtcc ggcgctcccc ccgcatcccc gagccggcag cgtgcgggga
     1321 cagcccgggc acggggaagg tggcacggga tcgctttcct ctgaacgctt ctcgctgctc
     1381 tttgagcctg cagacacctg gggggatacg gggaaaaagc taggctgaaa gaggatctta
     1441 gaatgacacg cgcctgggag ctcacgggga cagccccccc ccaaagcccc cagggatgta
     1501 attacgtccc tcccccgcta gggggcagca gcgagccgcc cggggctccg ctccggtccg
     1561 gcgctccccc cgcatccccg agccggcagc gtgcggggac agcccgggca cggggaaggt
     1621 ggcacgggat cgctttcctc tgaacgcttc tcgctgctct ttgagcctgc agacacctgg
     1681 ggggatacgg ggaaaaagct ggatcataac tcgacgtgca tactttaatt ggctccggtg
     1741 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgtgg ggaggggtcg
     1801 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
     1861 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
     1921 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
     1981 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
     2041 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
     2101 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
     2161 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
     2221 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
     2281 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
     2341 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
     2401 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
     2461 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     2521 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     2581 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     2641 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     2701 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     2761 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     2821 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     2881 gtttttttct tccatttcag gtggatccca agggcagata agtttgtaca aaaaagagag
     2941 gtgccaccat gagccccaag aagaagagaa aggtgtacct cagtaagatc atcatcgccc
     3001 gcgcttggtc ccgtgacctg taccaactgc accaagagct ctggcacctc ttccccaaca
     3061 ggccagatgc cgctagagac ttcctgttcc acgtggagaa gcgtaacacc cccgaagggt
     3121 gccacgtgct gttgcagagt gcccagatgc cagtgagtac cgctgttgcc actgtcatca
     3181 agactaaaca agttgaattc caactgcaag tgggcgtccc tctgtatttc cgcctcaggg
     3241 ccaaccccat caaaaccatc ctggacaacc agaagcggct ggatagcaaa ggtaatatca
     3301 agagatgccg cgtgcctctg atcaaggagg ccgagcagat cgcttggctg caacgcaagc
     3361 tgggtaacgc cgcgagagtg gaagatgtgc acccaatctc cgagcgcccg cagtatttct
     3421 ccggggaggg gaagaacggc aaaattcaga ctgtctgctt cgagggggtg ctcactatta
     3481 acgacgcccc tgctctgatc gacctcctgc agcagggcat tgggcccgcg aagagcatgg
     3541 gatgcggatt gttgagcctg gcacccctgt gagcttccac tggattgtac aattaccaac
     3601 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtgc gctggtgcgt
     3661 ttttatgctt gtagtattgt ataatgtttt taagatcctt aattaatagg gataacaggg
     3721 taatatcttg ccctgcacga ataccagcga ccccttgccc aaatacttgc cgtgggcctc
     3781 ggcctgagag ccaaaacact tgatgcggaa gaagtcggtg cgctcctgct tgtcgccggc
     3841 atcgttgcgc cacatctagg atctgccagg aaccgtaaaa aggccgcgtt gctggcgttt
     3901 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     3961 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4021 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4081 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4141 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4201 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4261 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4321 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4381 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4441 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     4501 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     4561 atga
//
