LOCUS Exported 7080 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Donor vector for 3' FLAG tag of human TGIF2. ACCESSION . VERSION . KEYWORDS pFETCh_TGIF2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7080) TITLE Mendenhall CRISPR tagging plasmids REFERENCE 2 (bases 1 to 7080) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7080 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 837..860 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 864..887 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 891..914 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 930..986 /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" CDS 999..1793 /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(1053..1072) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 1663..1682 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(2582..2599) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(2582..2598) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2591..2613) /label=M13/pUC Forward /note="In lacZ gene" CDS complement(2817..3632) /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind 3540..3559 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 3624..3643 /label=pENTR-R /note="pENTR vectors, reverse primer" rep_origin 4012..4600 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4501..4520 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 4754..4771 /label=L4440 /note="L4440 vector, forward primer" protein_bind 5071..5092 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5107..5137 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5145..5161 /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)." primer_bind 5150..5172 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 5169..5185 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 5169..5185 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" promoter 5362..5564 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 5515..5535 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 5561..5585 /label=LNCX /note="Human CMV promoter, forward primer" CDS 5661..6791 /codon_start=1 /product="herpes simplex virus thymidine kinase" /label=HSV TK /note="confers sensitivity to nucleoside analogs such as acyclovir or ganciclovir" /translation="MASYPCHQHASAFDQAARSRGHSNRRTALRPRRQQEATEVRLEQK MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWQVLGASETIANIYTTQ HRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGSSHAPPPALTLIFDR HPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGALPEDRHIDRLAKRQR PGERLDLAMLAAIRRVYGLLANTVRYLQGGGSWREDWGQLSGTAVPPQGAEPQSNAGPR PHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRPMHVFILDYDQSPAGCRDAL LQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN" ORIGIN 1 atcatagctc attgcaggct ccacctcttg ggctccagct atcttccgat ctcaggctcc 61 caagtagttg ggaccacagg ggtgcaccac tacacctggc taacttcttt tgtagaaaca 121 gggtctccct gtgttgccca ggctggtctt gaactcctgg gcttaagcaa tcctcccgcc 181 ttggccaccc aaagtgttgg gattacaggt gtgagccact gtgcccagcc catagctgtt 241 ttagattaag caaattgatc ggtcttgtat attcattcca gatatgtaac tggttcatca 301 atgcccggcg gcggcttctc ccagacatgc ttcggaagga tggcaaagac cctaatcagt 361 ttaccatttc ccgccgcggg ggtaaggcct cagatgtggc cctcccccgt ggcagcagcc 421 cctcagtgct ggctgtgtct gtcccagccc ccaccaatgt gctctccctg tctgtgtgct 481 ccatgccgct tcactcaggc cagggggaaa agccagcagc ccctttccca cgtggggagc 541 tggagtctcc caagcccctg gtgacccctg gtagcacact tactctgctg accagggctg 601 aggctggaag ccccacaggt ggactcttca acacgccacc acccacaccc ccagagcagg 661 acaaagagga cttcagcagc ttccagctgc tggtggaggt ggcgctacag agggctgctg 721 agatggagct tcagaagcag caggacccat cactcccatt actgcacact cccatccctt 781 tagtctctga aaatccccaa gggagcggag gaggttccgg tggaggtggt tctggagatt 841 acaaggatga cgacgataag ggcgattaca aggatgacga cgataaggga gattacaagg 901 atgacgacga taaggtttca ggaagcggag ctactaactt cagcctgctg aagcaggctg 961 gagacgtgga ggagaaccct ggacctggat cgtttcgcat gattgaacaa gatggattgc 1021 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga 1081 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt 1141 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat 1201 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg 1261 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg 1321 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc 1381 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga 1441 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag 1501 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc 1561 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg 1621 actgtggccg gctgggtgtg gccgaccgct atcaggacat agcgttggct acccgtgata 1681 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg 1741 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgattcgaac 1801 atcgaagggt gctgaaggct ccagccagct gtcctgggtt tccgttttgg ttccctttca 1861 tacagagggt tttctatgga tcactgccaa acattgggat catctcctct gtccagaggt 1921 cttcaacagg aagatgccag ctggcaccac tgcactgtga tgggggccct ctcctctgct 1981 gactctgccg tttctccagg cctccgctca gtgatgagac caagagatcg gagacaagca 2041 tggtgctgct gcttctgctg cttctccaga aaatccctgg gacacctttg ttccagcctg 2101 gtttcctggg ctgggctcag gaaagctgcc aaattcagtc ctatgttggg tccaagctgc 2161 ccctgtgctg tttctgtcaa gccaggtgtg gacattccaa gttcatatgc gtgaacaaaa 2221 gaaaagagga acccagtgga tgtaacagaa ccgactccag ttgaatgttt agatttttgc 2281 taaactgttt tctttttccc ttttttgctg tggtttgcat tcacggcagt agttagccca 2341 ggtgtgggga acgagagtgc actgcatgat agcgttctgg tgagctggga aggacccacc 2401 actgccactg aggattgttt tggaagaaag gaatattttt atcttgggga ccagctaagt 2461 ctctgcagta gtgtgaaatt ccaaatggtt gttttatcat tggtttggtt taccaaaaaa 2521 aaggcagggc ttggaaagtc ctctccactg actgtagcct ccaattcact ggccgtcgtt 2581 cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc 2641 gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc 2701 gcccttccca acagttgcgc agcctgaatg gcgaatgcga tttattcaac aaagccgccg 2761 tcccgtcaag tcagcgtaat gctctgccag tgttacaacc aattaaccaa ttctgattag 2821 aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt atcaatacca 2881 tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca gttccatagg 2941 atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat acaacctatt 3001 aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt gacgactgaa 3061 tccggtgaga atggcaaaag cttatgcatt tctttccaga cttgttcaac aggccagcca 3121 ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg tgattgcgcc 3181 tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg aatcgaatgc 3241 aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc aggatattct 3301 tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca tgcatcatca 3361 ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag ccagtttagt 3421 ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt cagaaacaac 3481 tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg cccgacatta 3541 tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa tcgcggcttc 3601 gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact gtttatgtaa 3661 gcagacagtt ttattgttca tgatgatata tttttatctt gtgcaatgta acatcagaga 3721 ttttgagaca caacgtggct ttgttgaata aatcgaactt ttgctgagtt gaaggatcag 3781 atcacgcatc ttcccgacaa cgcagaccgt tccgtggcaa agcaaaagtt caaaatcacc 3841 aactggtcca cctacaacaa agctctcatc aaccgtggct ccctcacttt ctggctggat 3901 gatggggcga ttcaggcctg gtatgagtca gcaacacctt cttcacgagg cagacctctc 3961 gacggagttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc 4021 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt 4081 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc 4141 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc 4201 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg 4261 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg 4321 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga 4381 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc 4441 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg 4501 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg 4561 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt 4621 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc 4681 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg 4741 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gaagctcgca cattcagcag 4801 cgtttttcag cgcgttttcg atcaacgttt caatgttggt atcaacacca ggtttaactt 4861 tgaacttatc ggcactgacg gttactgatt ttgaactttt gctttgccac ggaacggtct 4921 gcgttgtcgg gaagatgcgt gatctgatcc ttcaactcag caaaagttcc caatacgcaa 4981 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga 5041 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc 5101 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca 5161 atttcacaca ggaaacagct atgaccatga ttacgccaag cttgcatgcc tgcaggtcga 5221 ctctagagga tccccgggta ccgagctcga attagcttgc atgcctgcag gtcaattccc 5281 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta 5341 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag 5401 cggtttgact cacggggatt tcaagtctcc accccattga cgtcaatggg agtttgtttt 5461 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 5521 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 5581 agatcgcctg gagacgccat ccacgctgtt ttgacctcca taggagacac cgggaccgat 5641 ccagcctggg gatcccgacc atggcttcgt acccctgcca tcaacacgcg tctgcgttcg 5701 accaggctgc gcgttctcgc ggccatagca accgacgtac ggcgttgcgc cctcgccggc 5761 agcaagaagc cacggaagtc cgcctggagc agaaaatgcc cacgctactg cgggtttata 5821 tagacggtcc tcacgggatg gggaaaacca ccaccacgca actgctggtg gccctgggtt 5881 cgcgcgacga tatcgtctac gtacccgagc cgatgactta ctggcaggtg ctgggggctt 5941 ccgagacaat cgcgaacatc tacaccacac aacaccgcct cgaccagggt gagatatcgg 6001 ccggggacgc ggcggtggta atgacaagcg cccagataac aatgggcatg ccttatgccg 6061 tgaccgacgc cgttctggct cctcatatcg ggggggaggc tgggagctca catgccccgc 6121 ccccggccct caccctcatc ttcgaccgcc atcccatcgc cgccctcctg tgctacccgg 6181 ccgcgcgata ccttatgggc agcatgaccc cccaggccgt gctggcgttc gtggccctca 6241 tcccgccgac cttgcccggc acaaacatcg tgttgggggc ccttccggag gacagacaca 6301 tcgaccgcct ggccaaacgc cagcgccccg gcgagcggct tgacctggct atgctggccg 6361 cgattcgccg cgtttacggg ctgcttgcca atacggtgcg gtatctgcag ggcggcgggt 6421 cgtggcggga ggattgggga cagctttcgg ggacggccgt gccgccccag ggtgccgagc 6481 cccagagcaa cgcgggccca cgaccccata tcggggacac gttatttacc ctgtttcggg 6541 cccccgagtt gctggccccc aacggcgacc tgtacaacgt gtttgcctgg gccttggacg 6601 tcttggccaa acgcctccgt cccatgcacg tctttatcct ggattacgac caatcgcccg 6661 ccggctgccg ggacgccctg ctgcaactta cctccgggat ggtccagacc cacgtcacca 6721 cccccggctc cataccgacg atctgcgacc tggcgcgcac gtttgcccgg gagatggggg 6781 aggctaactg aggatcccac gtcactattg tatactctat attatactct atgttatact 6841 ctgtaatcct actcaataaa cgtgtcacgc ctgtgaaacc gtactaagtc tcccgtgtat 6901 tcttatcacc atcaggtgac atcctcgccc aggctgtcaa tcatgccggt atcgattcca 6961 gtagcaccgg ccccacgctg acaacccact cttgcagcgt tagcagcgcc cctcttaaca 7021 agccgacccc caccagcgtc gcggttacta acactcctct ccccgacctg cagcccagct //