LOCUS       sequence-275473-        7197 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPR donor plasmid to create GFP fusion proteins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7197)
  TITLE     ENCODE collection: CRISPR constructs
REFERENCE   2  (bases 1 to 7197)
  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: pARID4B-donor
FEATURES             Location/Qualifiers
     source          1..7197
                     /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             3669..3689
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3702..3746
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             3786..3809
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(3837..3839,3840..3842,3843..4556)
                     /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(3882..3903)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4143..4162)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    join(4594..5167,5168..5170,5171..5179)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4760..4777)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4987..5006
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5261..6064
                     /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(5324..5343)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5934..5953
                     /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 tagaagaaca 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 tcggtaccca ttttgggagg ctgagatggg cggatcatga
     2701 ggtcaggaga tcgagaccat cctggctaac atggtgaaac cccatctcta ctaaaaatac
     2761 aaaaaaatta gccgggcgtg gtggcgggcg cttatagtcc cagctactgg gggggctgag
     2821 gcaggagaat ggcgtgaact caggaggcag aacttttagt gagccaagat cgagccactg
     2881 cactccagcc tgggcgacag acagagcgag actttgtctc aaaaaacaaa caaacaaaaa
     2941 aaaacgaaaa aaggaaacag tgagagactt cctctgatga aaataactaa tgtgttattt
     3001 gctttgtaaa cctagtggcg gagaatgcaa cagttgagct atgcactgtt ttgatccaac
     3061 ttgaagaagc aattaagcct cccactcttg tcaccattta catgtacaag aaaactccta
     3121 agtacagaaa gatggaggga ttaggagggg acaaatgatt tttggatgga ttgcagattt
     3181 ttcctgtttt gatacttgtt tcactgttac aaaaagtgta ttctgtattt tatttctgtg
     3241 tcttgtagac taggcacagt tctttccctc tttgacccac gcaggagcct tccctggtct
     3301 gtccttttca ttacttctgt agttgggcac tgtctagctt cttgagctac actagctttt
     3361 cctttcttca catcgtagaa ctgtgagaat tgaccactgt tgttgtaagc taaatgattt
     3421 tggacaatat agcggaattt tagttcagag gactagtatt ttctgtctat tgttagacat
     3481 aaatttttat taagctcttg gtttggtctc ccttttcctt aggtgctgct acatcctcat
     3541 cctcctcttc accttcatcc agttccataa cagctgctgt tatgttaact ttagctgaac
     3601 cgtcaatgtc cagcgcatca caaaatggaa tgtcagttga gtgcaggggg attccaacta
     3661 ctgcaagcga gaatctttat tttcagggcg ccgccaaatt caaagaaacc gctgctgcta
     3721 aattcgaacg ccagcacatg gacagcggag gtggaggttc gagcggtccc tcgggttcgt
     3781 cgagcctgga agttctgttc caggggcccc tgtcgtcgag cggtccctcg ggttcgatgg
     3841 tgagcaaagg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
     3901 acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca
     3961 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
     4021 tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc
     4081 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca
     4141 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
     4201 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
     4261 tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca
     4321 tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc
     4381 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     4441 tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctta
     4501 aggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtaatgat
     4561 ctagagtcga gttaattaag aattccgccc ccccccctct ccctcccccc ccctaacgtt
     4621 actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt attttccacc
     4681 atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt cttgacgagc
     4741 attcctaggg gtctttcccc tctcgccaaa ggaatgcagg gtctgttgaa tgtcgtgaag
     4801 gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg
     4861 cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg tgtataagat
     4921 acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt tgtggaaaga
     4981 gtcaaatggc tctcctcaag cgtattcaac aaggggctga aggatgccca gaaggtaccc
     5041 cattgtatgg gatctgatct ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg
     5101 ttaaaaaaac gtctaggccc cccgaaccac ggggacgtgg ttttcctttg aaaaacacga
     5161 tgataatatg gccacaacct gttacattgc acaagataaa aatatatcat cacgaacagt
     5221 aaaactgtct gcttacataa acagtaatac aaggggtgtt atgggatcgg ccattgaaca
     5281 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     5341 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     5401 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc
     5461 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     5521 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     5581 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     5641 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     5701 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     5761 gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct
     5821 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     5881 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     5941 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     6001 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     6061 ctgagtccag caggacttgc taaagcactt tgcacttaat ggctgttgag ggccactttt
     6121 tttttatact gcacagtggc acaaaaaaat atcagacaag cactatttta tatttaaaaa
     6181 ttgtttcttg acaagctgac ttggcactta agtgcacttt tttatgaaga aaaagtacaa
     6241 tgaactgctt ttcctcaagc aataattgtt tccaacttgt ctgggaattg tgtgtctggt
     6301 aactggaagg ccttccactg tggcaaatgg aggcttttca ctgcctgtag agacaataca
     6361 gtaagcatag ttaaggggtg ggtcagaaca tgttaagata acttactgta tatgtattcc
     6421 cttgtatttt gttaaagctg gaacatttga tatttttcca tttatttatg aaaaaatatg
     6481 aacctatttt catttgtaca aggtaattgt tttttaaagc aagtcacctt agggtggctt
     6541 taattgtata agtcaagcac atgtaataaa ttcaaaacct gcagttaaca ggatattaga
     6601 catcaatcct ggtaaccaaa tattaaagat tctctttaaa aaagactgaa catgtttaca
     6661 ggtttgaatt aggctaaaag gtcttgcagt ggcttttcat ggcccttcaa attggaatgg
     6721 aactactgta ctttgccatt tttctataaa tcagtatttt tttttaattt tgatatacat
     6781 tgtgtgaaaa aagaaaatgg ctaataaact gtattaaatc ttaaacaatg tataaagatt
     6841 gtacttagcc agttcaaagt gtatatttat tcataatgaa ttataacagt tatatttttg
     6901 tgttttcttg taaatgtttc ttttccctta aatacagata attcatttgt attgcttatt
     6961 ttattatgag ctacaacaaa aggacttcag gaacaagtaa tgtattagta tggttcaaga
     7021 ttgttgatag gaactgtctc aaaaggatgg tggttatttt aaatataaat agctaatggg
     7081 ggtggtaggc ctataaaatt aaatgccttg tataaaatcc aaaatgaatg caaaattgtt
     7141 ttcacttgta ttgactttat gttgtatgat tccaatctct gttctgtttg gcacttg
//
