LOCUS Exported 6833 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION CRISPR donor plasmid to tag human transcription factor NR2F6 with GFP. ACCESSION . VERSION . KEYWORDS pNR2F6-donor SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6833) TITLE White lab ENCODE plasmids to tag human transcription factors with GFP REFERENCE 2 (bases 1 to 6833) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6833 /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 3573..3593 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" CDS 3606..3650 /codon_start=1 /product="affinity and epitope tag derived from pancreatic ribonuclease A" /label=S-Tag /translation="KETAAAKFERQHMDS" CDS 3690..3713 /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /label=HRV 3C site /translation="LEVLFQGP" CDS 3741..4460 /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(3786..3807) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(4047..4066) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" misc_feature 4498..5083 /label=IRES2 /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(4664..4681) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 4891..4910 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 5165..5968 /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(5228..5247) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 5838..5857 /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 aagagatcct cccacctcag cctcttgagg 2701 agctgggact acaagcgtgt gccatcacac tggactaatt tttatttttt atttattttg 2761 agacagggtc tccctctgtt gcccaggctg gagtgcagtg gcacgatctc ggctcactgc 2821 aacctccgcc tcccaggctc aagcgatcct cccattagcc ttccgagtag ctggaactgc 2881 aggcacgcac taccatgcct ggctaattat tcttattttt agtagagacg gaatatcatc 2941 atgttggcca ggctggtctt gaactcctga cctcaagtga tccacctgcc tcagcctccc 3001 aaagtgctgg gattacaggc gtgagctact gtgcccagcc ccagctgatt tgtttttgca 3061 gagaagggat catactatgt tgccctggct ggtctcaaat cctggcctta aatgatgctt 3121 ctgccttggc ctcccaaagt gctgggactg tggccgtgag ccatcatgcc cggccccctg 3181 ttcttaccca cagtggcgag tggtgtcccc tagtcacccc tgtgggctct gtgttctcta 3241 gctgcctccc acctgtctga cttgcctttg tccccctaga cgcctgtggc ctctcagacc 3301 cggcccacgt tgagagcctg caggagaagg cgcaggtggc cctcaccgag tatgtgcggg 3361 cgcagtaccc gtcccagccc cagcgcttcg ggcgcctgct gctgcggctc cccgccctgc 3421 gcgcggtccc tgcctccctc atctcccagc tgttcttcat gcgcctggtg gggaagacgc 3481 ccattgagac actgatcaga gacatgctgc tgtcggggag taccttcaac tggccctacg 3541 gctcgggcca ggggattcca actactgcaa gcgagaatct ttattttcag ggcgccgcca 3601 aattcaaaga aaccgctgct gctaaattcg aacgccagca catggacagc ggaggtggag 3661 gttcgagcgg tccctcgggt tcgtcgagcc tggaagttct gttccagggg cccctgtcgt 3721 cgagcggtcc ctcgggttcg atggtgagca aaggcgagga gctgttcacc ggggtggtgc 3781 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg 3841 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc 3901 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc 3961 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg 4021 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga 4081 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg 4141 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca 4201 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg 4261 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg 4321 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg 4381 agaagcgcga tcacatggtc cttaaggagt tcgtgaccgc cgccgggatc actctcggca 4441 tggacgagct gtacaagtaa tgatctagag tcgagttaat taagaattcc gccccccccc 4501 ctctccctcc ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg 4561 tttgtctata tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa 4621 cctggccctg tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg 4681 cagggtctgt tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca 4741 acgtctgtag cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc 4801 ggccaaaagc cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt 4861 gtgagttgga tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg 4921 ctgaaggatg cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca 4981 tgctttacat gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac 5041 gtggttttcc tttgaaaaac acgatgataa tatggccaca acctgttaca ttgcacaaga 5101 taaaaatata tcatcacgaa cagtaaaact gtctgcttac ataaacagta atacaagggg 5161 tgttatggga tcggccattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt 5221 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt 5281 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc 5341 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc 5401 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga 5461 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat 5521 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca 5581 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga 5641 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc 5701 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat 5761 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga 5821 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg 5881 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt 5941 ctatcgcctt cttgacgagt tcttctgagt ctgaccatga cggggccacg tgtgctgtgg 6001 ccaggcctgc agacagacct caagggacag ggaatgctga ggcctcgagg ggcctcccgg 6061 ggcccaggac tctggcttct ctcctcagac ttctattttt taaagactgt gaaatgtttg 6121 tcttttctgt tttttaaatg atcatgaaac caaaaagaga ctgatcatcc aggcctcagc 6181 ctcatcctcc ccaggacccc tgtccaggat ggagggtcca atcctaggac agccttgttc 6241 ctcagcaccc ctagcatgaa cttgtgggat ggtggggttg gcttccctgg catgatggac 6301 aaaggcctgg cgtcggccag aggggctgct ccagtgggca ggggtagcta gcgtgtgcca 6361 ggcagatcct ctggacacgt aacctatgtc agacactaca tgatgactca aggccaataa 6421 taaagacatt tcctacctgc acagcctccc tggtctttat ttttattttt tatttttgag 6481 acggagtcct acttcgtcac ccaggctgga gtgcagtggc gtgatcttgg ctcactgtaa 6541 cctctgcctc acaggttcaa gcgattctcc tgcctcagcc tccccagtag ctggaactac 6601 aggcacccac cacgatgccc agctaatttt atttttggta gagatggggt ttcaccatgt 6661 tgaccaggtt ggtctcgaac tcctgacctc aggtgatgca cctgaggcct ccaaaagtgg 6721 attacaagcg tgagccactg tgcctagcct cacttacttt cttttggtca accttaatct 6781 tttttttttt tttttttttt ttttttgaga tggagtttcg ctgtttcacc tgg //