LOCUS Exported 7486 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION CRISPR donor plasmid to tag human transcription factor TCF15 with GFP. ACCESSION . VERSION . KEYWORDS pTCF15-donor SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7486) TITLE White lab ENCODE plasmids to tag human transcription factors with GFP REFERENCE 2 (bases 1 to 7486) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7486 /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 3889..3909 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" CDS 3922..3966 /codon_start=1 /product="affinity and epitope tag derived from pancreatic ribonuclease A" /label=S-Tag /translation="KETAAAKFERQHMDS" CDS 4006..4029 /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /label=HRV 3C site /translation="LEVLFQGP" CDS 4057..4776 /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(4102..4123) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(4363..4382) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" misc_feature 4814..5399 /label=IRES2 /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(4980..4997) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 5207..5226 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 5481..6284 /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(5544..5563) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 6154..6173 /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 tcggtaccct ctggcttacg gtgaggatga aacccatgtg 2701 tccagtctga aaatatcaat gagaagagct ggtagagaca gcccctgtaa ggactgatgg 2761 tttcagggac tgccgccatt agcccatctt ccaatggctg ccacttaatt caggcggcct 2821 tgattggtgc ctgctccaag tcaggtcctg ggccaagaga gcccaggctg gaaatgggac 2881 cacagagtga agaactcctt ttccctacca ctccctgaga tcccagtcca gatggggaga 2941 caaaggagag accctgcctt gtccttctga gattcatccc catggaatgt ccccagaaag 3001 gcaggaagtg ctaggcaatt tcaggaacag agaaaccttt ccagggagac aagacctgag 3061 agcacattcc tgggaaaggg gacgccctga gggaagtcaa ggaggccaga aaccgagtcc 3121 ccttttaaag gtggacgagt gagccgctag gcatgcagtc aggcaggggg aggaccactg 3181 tggggtgtgc ttgttttgcc cgtgtaaaat attatgcttt aaaatatatt ggattttttt 3241 taagcaacag atcatatggg aaagaagtag aaaaggtaaa aaagtatagc gtgaatacaa 3301 aaattagccg ggtgtggcgg tgggcccctg tagtccccgc tactcgggag gatgaggcag 3361 gagaatcact tgaacccggg aggcagaggt tgcagtgagc tgagattgtg ccactgcatt 3421 ccagcctggc aacagagcaa gactccatct caaaaaacag tatagcatga gagagcctcc 3481 ctcccatcct ctctccatca tcaaattcct ctctgaagaa acccactcac aatttcccgt 3541 gtgtgcttct ccccagcaat cacagcacac aggcgcattc agcctgcccc ttgcttttgt 3601 ccctaaacat tgtcatttgg cagcagtctg tatcagtcca gaaagggttt cttctgcttt 3661 ctgttgcaga gaacttgatc gtgtggctga actgtgattt attgatcagg atccctcttg 3721 agggatacag aggttagttc cagtgttcac actggcctaa tcgaacctct ttatactctt 3781 tctccccctc tgcagggtgg ccgtcgtgac ctggggggca gctgcttgaa ggtgaggggg 3841 gtggcccccc ttcgagggcc acggagaggg attccaacta ctgcaagcga gaatctttat 3901 tttcagggcg ccgccaaatt caaagaaacc gctgctgcta aattcgaacg ccagcacatg 3961 gacagcggag gtggaggttc gagcggtccc tcgggttcgt cgagcctgga agttctgttc 4021 caggggcccc tgtcgtcgag cggtccctcg ggttcgatgg tgagcaaagg cgaggagctg 4081 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc 4141 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc 4201 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc 4261 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc 4321 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag 4381 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc 4441 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc 4501 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc 4561 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc 4621 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg 4681 agcaaagacc ccaacgagaa gcgcgatcac atggtcctta aggagttcgt gaccgccgcc 4741 gggatcactc tcggcatgga cgagctgtac aagtaatgat ctagagtcga gttaattaag 4801 aattccgccc ccccccctct ccctcccccc ccctaacgtt actggccgaa gccgcttgga 4861 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa 4921 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc 4981 tctcgccaaa ggaatgcagg gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc 5041 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg 5101 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca 5161 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag 5221 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct 5281 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc 5341 cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataatatg gccacaacct 5401 gttacattgc acaagataaa aatatatcat cacgaacagt aaaactgtct gcttacataa 5461 acagtaatac aaggggtgtt atgggatcgg ccattgaaca agatggattg cacgcaggtt 5521 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct 5581 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga 5641 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg 5701 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact 5761 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg 5821 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct 5881 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg 5941 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt 6001 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg 6061 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc 6121 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag 6181 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt 6241 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagtcgcc tggaccctgg 6301 agaaggaggc caggagccag ccactggctg gacagggaag aagaccccag gagccaagcc 6361 caccccttct ttgtgtaggg accgggggac catggcctgt tccgggacac tctgggcagg 6421 gccctcggga catctccacc cgatcctgga gagctgtgag gatccattca gcctgcccag 6481 ctctggctgg tcagagacaa ggcagaactt ttggaaaaac aaagactgtt ggtgacaggg 6541 tgtgtgtgta tctgtgcgtg agtgtgagtg tgtgtgagag agaattggtg agtttaaaat 6601 aaaagctatt tttaaataaa agacgtcgtt ctgagctgag gagagtcccg tggacatcgg 6661 aggtcagagg tgggccctcc cctggaccta gagagaggca cagatgttct gggggctgcc 6721 ctgagccctg gaagggtggg ggtggggcgg gggaaggcag gcagagcccc ctctccgggc 6781 gccataacca aactttcctc agtctgctgt ttatcagatg ttcttaacaa ctgttaggtg 6841 tctgatatgg ttggtaaaga aatcttaatt ttacttattt ttgaataagg cctgtgttca 6901 tttcattcac atggttcaaa actcagaagg taccagaggg tatgtgttga aaagtctccg 6961 tgcccctgtc ctccagccac cagctctcct ctctgcaagc agcctagggc accagtttcc 7021 tgtgatgtcc cattcactat gaatgatcag gcaaatacac aaatgacagc acatcgtaca 7081 cacttttggc tccttctttt tttccttaat ctagcttggt atcatcccaa gtcagtacct 7141 aaagggtatt tcatttcttc ctgagctgct tattattcca ttttgtgaac atgccatcct 7201 ttaacccatc cctctttgat gcgatcacca acaagccatg tggttttcca caagtgcaaa 7261 tgagtgcatg ctaaagggga atattcttac ccctgtgtta gctgaaatgc tttagaagac 7321 cgatgacacc cagaggcacg tggcagggtg acaccctgag gtccgtccac ctttgaagcc 7381 cagagcagct ccttccttgc cctggagatg ccccaagtgg aaagccagca cctgttaccc 7441 caattacact cccaggagcc ctgggaggtg agagctggtg tcttcc //