LOCUS       sequence-211742-        7124 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPR donor plasmid to tag human transcription factor E2F5 with 
            GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7124)
  TITLE     White lab ENCODE plasmids to tag human transcription factors with 
            GFP
REFERENCE   2  (bases 1 to 7124)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pE2F5-donor
FEATURES             Location/Qualifiers
     source          1..7124
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(34..50)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(34..50)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(47..69)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    58..74
                     /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(join(82..88,89..106,107..112))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    127..148
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(265..282)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(436..1024)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(516..535)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1195..1986,1987..2055))
                     /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     1818..1837
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2056..2160)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2228..2246
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2284..2306)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2406..2425
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2619..2641
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2633..2650
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2634..2650
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3641..3661
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3674..3718
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             3758..3781
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(3809..3811,3812..3814,3815..4528)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLK
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3854..3875)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4115..4134)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    join(4566..5139,5140..5142,5143..5151)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4732..4749)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4959..4978
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5233..6036
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5296..5315)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5906..5925
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 gacctgcagg catgcaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
       61 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
      121 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
      181 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
      241 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
      301 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
      361 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
      421 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
      481 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
      541 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
      601 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
      661 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
      721 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
      781 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
      841 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
      901 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
      961 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     1021 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     1081 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     1141 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     1201 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     1261 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     1321 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     1381 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     1441 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     1501 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     1561 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     1621 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     1681 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     1741 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     1801 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     1861 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     1921 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     1981 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     2041 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     2101 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     2161 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     2221 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     2281 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     2341 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     2401 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
     2461 cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg
     2521 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     2581 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     2641 gacggccagt gaattcgagc tcggtacccg tctccctctg tcatccaggc tggagtgtca
     2701 tccaggctgg agtgcagtgg catgatagct cactgcaacc tccacctcct aggttcaaat
     2761 gattctcctg gctcagcctc ccgagtagct gggatgacag gcacctgcca ccacacctgg
     2821 ctactttttg tatttttagt agagacgggg tttcaccatg ttggccaggc tggtctcaaa
     2881 ctcctggcct caggtgatcc acctgcctca gcctcccaaa gtgttgggat tacaggcatg
     2941 agccactgca cctggcctcg ttagtgcctt ttagttatat taccttagtg ataagttcaa
     3001 ccaataaaaa atttaatttc taggccgggc acaatggctc atgcctgtaa tcccagcact
     3061 ttgggaggcc aaggcgggcg aatcacgtca ggagatcgag accatcctgg ctaacatggt
     3121 gaaaccccat ctctataaaa aaataaaaaa aatgagccgg tcgtggtggc aggcacctgt
     3181 agtcccagct actcgggagg ctgaggcagg agaatggcgt gaacccggga gtcgggagct
     3241 tgcagtgggc cgagatcgag ctactgcact ccagcctgga cgaaacaggg agactccatc
     3301 ttaaaaaaaa aaaaaaaaaa agaaaaaatt aacttctgtt tctcaatcta atatcaatat
     3361 tcaatgtaca aattgttact aaatgagaat atttcaaaat acacattaca caagacttta
     3421 atttggaatg atgtatttga gagtattttg ttaaagatca ttaaccgtat gtagaaacca
     3481 tctttaacac taaatttgtt ttttgttcgc agtgtttcct ctcttaaggc tttctcctac
     3541 cccggcagat gactacaact ttaatttaga tgataacgaa ggagtttgtg atctgtttga
     3601 tgtccagata ctaaattatg ggattccaac tactgcaagc gagaatcttt attttcaggg
     3661 cgccgccaaa ttcaaagaaa ccgctgctgc taaattcgaa cgccagcaca tggacagcgg
     3721 aggtggaggt tcgagcggtc cctcgggttc gtcgagcctg gaagttctgt tccaggggcc
     3781 cctgtcgtcg agcggtccct cgggttcgat ggtgagcaaa ggcgaggagc tgttcaccgg
     3841 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     3901 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     3961 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
     4021 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     4081 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     4141 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     4201 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     4261 ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa
     4321 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     4381 cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga
     4441 ccccaacgag aagcgcgatc acatggtcct taaggagttc gtgaccgccg ccgggatcac
     4501 tctcggcatg gacgagctgt acaagtaatg atctagagtc gagttaatta agaattccgc
     4561 ccccccccct ctccctcccc ccccctaacg ttactggccg aagccgcttg gaataaggcc
     4621 ggtgtgcgtt tgtctatatg ttattttcca ccatattgcc gtcttttggc aatgtgaggg
     4681 cccggaaacc tggccctgtc ttcttgacga gcattcctag gggtctttcc cctctcgcca
     4741 aaggaatgca gggtctgttg aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa
     4801 gacaaacaac gtctgtagcg accctttgca ggcagcggaa ccccccacct ggcgacaggt
     4861 gcctctgcgg ccaaaagcca cgtgtataag atacacctgc aaaggcggca caaccccagt
     4921 gccacgttgt gagttggata gttgtggaaa gagtcaaatg gctctcctca agcgtattca
     4981 acaaggggct gaaggatgcc cagaaggtac cccattgtat gggatctgat ctggggcctc
     5041 ggtgcacatg ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc
     5101 acggggacgt ggttttcctt tgaaaaacac gatgataata tggccacaac ctgttacatt
     5161 gcacaagata aaaatatatc atcacgaaca gtaaaactgt ctgcttacat aaacagtaat
     5221 acaaggggtg ttatgggatc ggccattgaa caagatggat tgcacgcagg ttctccggcc
     5281 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     5341 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     5401 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     5461 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     5521 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     5581 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     5641 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     5701 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     5761 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     5821 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     5881 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     5941 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     6001 atcgccttct atcgccttct tgacgagttc ttctgagtct agattccatg gaaacttggg
     6061 actgttatct acctctaact gtgtaacatt ttagacttct taataaccta aatatttaaa
     6121 ataatgaatg taacaccttt tttagttcac tgattctgaa gtgttcttcc ctaatacttt
     6181 ctttacttca caaaacttca accataaaaa caaagggctc tgattgcttt aggggataag
     6241 tgatttaata tccacaaacg tccccactcc caaaagtaac tatattctgg atttcaactt
     6301 ttcttctaat tgtgaatcct tctgtttttt cttcttaagg aggaaagtta aaggacacta
     6361 caggtcatca aaaacaagtt ggccaaggac tcattacttg tcttatattt ttactgccac
     6421 taaactgcct gtatttctgt atgtccttct atccaaacag acgttcactg ccacttgtaa
     6481 agtgaaggat gtaaacgagg atatataact gtttcagtga acagattttg tgaagtgcct
     6541 tctgttttag cactttaagt ttatcacatt ttgttgactt ctgacattcc actttcctag
     6601 gttataggaa agatctgttt atgtagtttg tttttaaaat gtgccaatgc ctgtacatta
     6661 acaagatttt taaaaataaa attgtataaa acattcaatt tattggtctt tgtggagaat
     6721 tagatgcatc accagtatat tacaacagag ccattaatct tgtagcttca tcaacattaa
     6781 ctggtttgct ttcatgacgc tgctgaggaa tctgaaagga gaaagtatta tatttaaaaa
     6841 cacagacaac aatagcagac aaatgactta tataagctgt tattcaattt aatttttcta
     6901 gtcatatctt tggaagtaga acaatctgta tattctgaca atgttttaga gtctgaatat
     6961 gctatttata ctatagagcc tagtactaga gagccaagta ctaaaaatta cactcaactt
     7021 acaggctttg aaaacttggg agtttaattt attagcctca atttgttagc cataaaaaaa
     7081 agcaaatgat ttctgcttac cagttctttc tgcagaggtt caag
//
