LOCUS Exported 4804 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Template for C-terminal PCR tagging of mammalian genes (Puromycin selection) with sfGFP. ACCESSION . VERSION . KEYWORDS pMaCTag-P06 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4804) AUTHORS Fueller J, Herbst K, Meurer M, Gubicza K, Kurtulmus B, Knopf JD, Kirrmaier D, Buchmuller BC, Pereira G, Lemberg MK, Knop M TITLE CRISPR-Cas12a-assisted PCR tagging of mammalian genes. JOURNAL J Cell Biol. 2020 Jun 1;219(6). pii: 151766. doi: 10.1083/jcb.201910210. PUBMED 32406907 REFERENCE 2 (bases 1 to 4804) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4804 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 1..19 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 1..18 /label=SP6 /note="SP6 promoter, forward primer" polyA_signal 801..922 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(838..857) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 892..911 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" protein_bind 929..962 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." promoter 969..1285 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 1136..1271 /label=SV40 ori /note="SV40 origin of replication" primer_bind 1198..1217 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" polyA_signal 1907..2131 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" promoter 2138..2378 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 2138..2158 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 2309..2328 /label=LKO.1 5' /note="Human U6 promoter, forward primer" protein_bind 2388..2421 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind complement(2458..2477) /label=T7 /note="T7 promoter, forward primer" promoter complement(2459..2477) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2564..2581) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(2735..3323) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2815..2834) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(3494..4354) /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 4117..4136 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(4355..4459) /gene="bla" /label=AmpR promoter primer_bind 4527..4545 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(4583..4605) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 4705..4724 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 atttaggtga cactatagaa cgcggccgcc agctgaagct tcgtacgtca ggtggaggag 61 gtagtggcgg aggcggatcc agcaagggcg aggagctgtt caccggcgtg gtgcccatcc 121 tggtggagct ggacggcgac gtgaacggcc acaagttcag cgtgcgcggc gagggcgagg 181 gcgacgccac caacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg 241 tgccctggcc caccctggtg accaccctga cctacggcgt gcagtgcttc agccgctacc 301 ccgaccacat gaagcgccac gacttcttca agagcgccat gcccgagggc tacgtgcagg 361 agcgcaccat cagcttcaag gacgacggca cctacaagac ccgcgccgag gtgaagttcg 421 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca 481 acatcctggg ccacaagctg gagtacaact tcaacagcca caacgtgtac atcaccgccg 541 acaagcagaa gaacggcatc aaggccaact tcaagatccg ccacaacgtg gaggacggca 601 gcgtgcagct ggccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc 661 tgcccgacaa ccactacctg agcacccaga gccgcctgag caaggacccc aacgagaagc 721 gcgaccacat ggtgctgctg gagttcgtga ccgccgccgg catcacccac ggcatggacg 781 agctgtacaa gtgaactagt aacttgttta ttgcagctta taatggttac aaataaagca 841 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt 901 ccaaactcat caatgtatct tagtcgatat aacttcgtat agcatacatt atacgaagtt 961 atgtcgacgg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca 1021 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat 1081 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc 1141 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat 1201 ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt 1261 ttttggaggc ctaggctttt gcaaagaatt cgccaccatg actgagtaca agcctactgt 1321 gcgtttagcc accagggatg atgtgccacg agccgtgagg acactagcag ccgcctttgc 1381 tgactaccca gctacaaggc acaccgtgga ccctgacaga cacattgaaa gagtcacaga 1441 acttcaggag ctgtttttaa ctagggtggg cctggatatt ggcaaagtct gggtggccga 1501 tgatggagca gcagttgcag tgtggaccac cccagagtct gttgaagctg gagctgtctt 1561 tgcagaaatt ggaccaagaa tggccgagtt atctggcagc aggctggctg ctcaacagca 1621 gatggaagga ttattagctc cccacagacc caaggagcct gcctggttcc tggcaacagt 1681 tggagtttct ccagaccatc agggcaaggg cttaggatct gctgtggtgc tgcctggtgt 1741 ggaagcagct gagagagctg gggtgccagc atttctggag acatcagctc ctagaaacct 1801 gcccttctat gaacggctgg gcttcacagt gactgctgat gtggaggtgc ctgagggccc 1861 cagaacctgg tgcatgacaa gaaaaccagg agcctgactg cagcgactgt gccttctagt 1921 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact 1981 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat 2041 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc 2101 aggcatgctg gggatgcggt gggctctatg gctcgaggag ggcctatttc ccatgattcc 2161 ttcatatttg catatacgat acaaggctgt tagagagata attggaatta atttgactgt 2221 aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt 2281 tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta 2341 tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccgata acttcgtata 2401 gcatacatta tacgaagtta tggtaccgat gcagctagcc cgcggatctg ccggtctccc 2461 tatagtgagt cgtattaatt tcgataagcc aggttaacct gcattaatga atcggccaac 2521 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc 2581 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt 2641 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg 2701 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg 2761 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat 2821 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta 2881 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct 2941 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc 3001 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa 3061 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg 3121 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag 3181 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt 3241 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta 3301 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc 3361 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca 3421 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa 3481 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat 3541 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct 3601 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt 3661 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat 3721 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta 3781 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg 3841 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt 3901 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg 3961 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg 4021 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc 4081 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa 4141 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac 4201 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt 4261 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg 4321 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa 4381 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata 4441 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca 4501 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc 4561 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt 4621 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg 4681 ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata 4741 tggacatatt gtcgttagaa cgcggctaca attaatacat aaccttatgt atcatacaca 4801 tacg //