LOCUS       Exported                7194 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  CRISPR donor plasmid to tag human transcription factor SPDEF with 
            GFP.
ACCESSION   .
VERSION     .
KEYWORDS    pSPDEF-donor
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7194)
  TITLE     White lab ENCODE plasmids to tag human transcription factors with 
            GFP
REFERENCE   2  (bases 1 to 7194)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7194
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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(82..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(1195..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             3233..3244
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             3874..3894
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3907..3951
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             3991..4014
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4042..4761
                     /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(4087..4108)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4348..4367)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    4799..5384
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4965..4982)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     5192..5211
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5466..6269
                     /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(5529..5548)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6139..6158
                     /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 tcggtacccc tgccaatgaa gccgggactg gcctcccatt
     2701 cccactcacc gccaccctcc ctggccaccc agcctcgacc agtgaggaga gctggaccga
     2761 cagcgaggtg gactcatcat gctccgggca gcccatccac ctgtggcagt tcctcaagga
     2821 gttgctactc aagccccaca gctatggccg cttcattagg tggctcaaca aggagaaggg
     2881 tgagcagggt gcccaagggg cgctgcaggg ccagtgagca tggaggttgt gggggcttct
     2941 gccaggcacg gtgcatgggg gcaggggcgg gcagaggggt gaagctgatc cccacctttg
     3001 tcctagatgc caaatatgca gctctcatgg gacagccctg gggtctgtgt gcaatctgtg
     3061 tccacatctg tctcctgcta gactctgtct cacttaggag ttttagtact gcacagcacc
     3121 tggaacgagc tccaaatcca ctctccctcc accatctcat attaattaat cctgccagag
     3181 agggctgcac aggccgaact gcctgagaat agcaaagagg ttgtttcagg cttgggaact
     3241 acagagacac ctgtaatggg gaagggatgc tcttgccaag tgagccgtgg gcactgctgg
     3301 gagccacaca ggactttgca ttaggtcatg caaaccccac acagaagcag gagttggcaa
     3361 gggccaggcc tgcagggccc cagggaaccc actaaggtac acaggctcac ctgtaccggg
     3421 aggtttgtga ctccagagta ttctctgcca ttcactgggg cctgtggcac ctgctttgca
     3481 cacgtctctt taacctcacc gcaactacac cttgggagta ggcatttggg cccccacacc
     3541 ccgccccacc cgagcagggg gaggcctctg cacggggtcc cttggttcgc cacgaggctg
     3601 acctggtggt ctgcgtcttt ctccttcttg ccacctcact cataccactg gctctctcgg
     3661 ggccctcctg ctctaggcat cttcaaaatt gaggactcag cccaggtggc ccggctgtgg
     3721 ggcatccgca agaaccgtcc cgccatgaac tacgacaagc tgagccgctc catccgccag
     3781 tattacaaga agggcatcat ccggaagcca gacatctccc agcgcctcgt ctaccagttc
     3841 gtgcacccca tcgggattcc aactactgca agcgagaatc tttattttca gggcgccgcc
     3901 aaattcaaag aaaccgctgc tgctaaattc gaacgccagc acatggacag cggaggtgga
     3961 ggttcgagcg gtccctcggg ttcgtcgagc ctggaagttc tgttccaggg gcccctgtcg
     4021 tcgagcggtc cctcgggttc gatggtgagc aaaggcgagg agctgttcac cggggtggtg
     4081 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
     4141 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     4201 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc
     4261 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     4321 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     4381 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     4441 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     4501 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     4561 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     4621 gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac
     4681 gagaagcgcg atcacatggt ccttaaggag ttcgtgaccg ccgccgggat cactctcggc
     4741 atggacgagc tgtacaagta atgatctaga gtcgagttaa ttaagaattc cgcccccccc
     4801 cctctccctc ccccccccta acgttactgg ccgaagccgc ttggaataag gccggtgtgc
     4861 gtttgtctat atgttatttt ccaccatatt gccgtctttt ggcaatgtga gggcccggaa
     4921 acctggccct gtcttcttga cgagcattcc taggggtctt tcccctctcg ccaaaggaat
     4981 gcagggtctg ttgaatgtcg tgaaggaagc agttcctctg gaagcttctt gaagacaaac
     5041 aacgtctgta gcgacccttt gcaggcagcg gaacccccca cctggcgaca ggtgcctctg
     5101 cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc agtgccacgt
     5161 tgtgagttgg atagttgtgg aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg
     5221 gctgaaggat gcccagaagg taccccattg tatgggatct gatctggggc ctcggtgcac
     5281 atgctttaca tgtgtttagt cgaggttaaa aaaacgtcta ggccccccga accacgggga
     5341 cgtggttttc ctttgaaaaa cacgatgata atatggccac aacctgttac attgcacaag
     5401 ataaaaatat atcatcacga acagtaaaac tgtctgctta cataaacagt aatacaaggg
     5461 gtgttatggg atcggccatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     5521 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     5581 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     5641 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     5701 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     5761 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     5821 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     5881 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     5941 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     6001 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     6061 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     6121 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     6181 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     6241 tctatcgcct tcttgacgag ttcttctgag tcgtgcctgg cccagggcct gaaacccgcc
     6301 ctcaggggcc tctctcctgc ctgccctgcc tcagccaggc cctgagatgg gggaaaacgg
     6361 gcagtctgct ctgctgctct gaccttccag agcccaaggt cagggagggg caaccaactg
     6421 ccccaggggg atatgggtcc tctggggcct tcgggaccct ggggcagggg tgcttcctcc
     6481 tcaggcccag ctgctcccct ggaggacaga gggagacagg gctgctcccc aacacctgcc
     6541 tctgacccca gcatttccag agcagagcct acagaagggc agtgactcga caaaggccac
     6601 aggcagtcca ggcctctctc tgctccatcc ccctgcctcc cattctgcac cacacctggc
     6661 atggtgcagg gagacatctg cacccctgag ttgggcagcc aggagtgccc ccgggaatgg
     6721 ataataaaga tactagagaa ctgacatgtc cccagtgtct gcgcagtgaa tggcacagcc
     6781 ccgaggtcat caatggtcaa atgcggctgt ggtcacaggg cctggagtag tcaagggagg
     6841 ttttctggag gaggtggctt tgggctgagc ctaaagggca agggataatt gattggtaga
     6901 tgcttctggg agagagatag tgaggcttca aaatcaagga ggggctagaa cagccacctc
     6961 atcttccctg tcgctgttcg ggctgagagg caggcggcaa ggccggtgct aaagaggtgc
     7021 tacatgcagg accggagtgg ggaaggcatc tttcgcatcc ttttgctctg ttgcagagca
     7081 gggctttgca gctaacaggg tggaagagag gttggttggc tcctgtttat gtttcctgaa
     7141 ggcctcagca gtgctgtctg tgaagatgtg agctcctgtg agtgggtgtg agac
//
