LOCUS       Exported                4814 bp ds-DNA     circular SYN 11-FEB-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 4814)
  AUTHORS   Baylor-NRI
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Dec 16, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4814
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     terminator      268..295
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      387..473
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     550..568
                     /label=NheI-KRAS(WT)-F
     protein_bind    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     670..761
                     /label=oJDW 1653 - BclI_KOZAK_V5_GATA1_FWD
     regulatory      675..684
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             684..725
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     misc_feature    726..740
                     /label=G2AG2 Linker
     CDS             741..1747
                     /codon_start=1
                     /label=hsGATA1
                     /translation="MEFPGLGSLGTSEPLPQFVDPALVSSTPESGVFFPSGPEGLDAAA
                     SSTAPSTATAAAAALAYYRDAEAYRHSPVFQVYPLLNCMEGIPGGSPYAGWAYGKTGLY
                     PASTVCPTREDSPPQAVEDLDGKGSTSFLETLKTERLSPDLLTLGPALPSSLPVPNSAY
                     GGPDFSSTFFSPTGSPLNSAAYSSPKLRGTLPLPPCEARECVNCGATATPLWRRDRTGH
                     YLCNACGLYHKMNGQNRPLIRPKKRLIVSKRAGTQCTNCQTTTTTLWRRNASGDPVCNA
                     CGLYYKLHHQHYCGGSAQLMRAQSMASRGGVVSFSSCSQNSGQPKSLGPRHPLA"
     primer_bind     1153..1172
                     /label=oJDW 1458
     primer_bind     complement(1730..1754)
                     
     CDS             1758..1814
                     /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"
     primer_bind     1790..1840
                     /label=oJDW 1017 (P2A_AvrII_H2A_FWD)
     primer_bind     1790..1840
                     /label=oJDW 1017
     regulatory      1816..1823
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1821..2204
                     /codon_start=1
                     /label=H2A.F/Z
                     /translation="MAGGKAGKDSGKAKTKAVSRSQRAGLQFPVGRIHRHLKTRTTSHG
                     RVGATAAVYSAAILEYLTAEVLELAGNASKDLKVKRITPRHLQLAIRGDEELDSLIKAT
                     IAGGGVIPHIHKSLIGKKGQQKTA"
     primer_bind     1821..1840
                     /label=oJDW 1370
     CDS             2211..2921
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     CDS             2211..2918
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2897..2918)
                     /label=oJDW 1371
     primer_bind     complement(2899..2959)
                     /label=oJDW 1018
     protein_bind    complement(2939..3038)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3039..3059)
                     /label=PmeI-HindIII-KRAS(WT)-R
     promoter        complement(3056..3074)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3079..3095)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3208..4017
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      4164..4752
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctgg atcagccacc atgggtaagc ctatccctaa ccctctcctc ggtctcgatt
      721 ctacgggtgg tgctggtggt atggagttcc ctggcctggg gtccctgggg acctcagagc
      781 ccctccccca gtttgtggat cctgctctgg tgtcctccac accagaatca ggggttttct
      841 tcccctctgg gcctgagggc ttggatgcag cagcttcctc cactgccccg agcacagcca
      901 ccgctgcagc tgcggcactg gcctactaca gggacgctga ggcctacaga cactccccag
      961 tctttcaggt gtacccattg ctcaactgta tggaggggat cccagggggc tcaccatatg
     1021 ccggctgggc ctacggcaag acggggctct accctgcctc aactgtgtgt cccacccgcg
     1081 aggactctcc tccccaggcc gtggaagatc tggatggaaa aggcagcacc agcttcctgg
     1141 agactttgaa gacagagcgg ctgagcccag acctcctgac cctgggacct gcactgcctt
     1201 catcactccc tgtccccaat agtgcttatg ggggccctga cttttccagt accttctttt
     1261 ctcccaccgg gagccccctc aattcagcag cctattcctc tcccaagctt cgtggaactc
     1321 tccccctgcc tccctgtgag gccagggagt gtgtgaactg cggagcaaca gccactccac
     1381 tgtggcggag ggacaggaca ggccactacc tatgcaacgc ctgcggcctc tatcacaaga
     1441 tgaatgggca gaacaggccc ctcatccggc ccaagaagcg cctgattgtc agtaaacggg
     1501 caggtactca gtgcaccaac tgccagacga ccaccacgac actgtggcgg agaaatgcca
     1561 gtggggatcc cgtgtgcaat gcctgcggcc tctactacaa gctacaccac cagcactact
     1621 gtggtggctc cgctcagctc atgagggcac agagcatggc ctccagagga ggggtggtgt
     1681 ccttctcctc ttgtagccag aattctggac aacccaagtc tctgggcccc aggcaccccc
     1741 tggcttgcgg taccggagcc actaacttct ccctgttgaa acaagcaggg gatgtcgaag
     1801 agaatcccgg gccacctagg atggcaggtg gaaaagcagg taaagacagt ggaaaagcca
     1861 agacgaaagc cgtctccaga tctcagagag ctggtctgca gttccctgta ggtcgtatcc
     1921 acaggcacct gaagacccga accaccagcc atggccgtgt cggagctaca gctgcggttt
     1981 acagtgctgc tatcctggag tatctcaccg ctgaggtgct ggagttggct ggaaacgctt
     2041 ctaaggatct gaaggtgaag cgaatcactc ccagacattt acagctggcc atccgcggag
     2101 acgaggagct ggactcgctt atcaaggcca ccattgctgg aggaggagtc attcctcaca
     2161 tccacaagtc tctgattggt aagaagggcc agcagaaaac cgcagggggg atggtgagca
     2221 agggcgagga ggacaacatg gccatcatca aggagttcat gcgcttcaag gtgcacatgg
     2281 agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg
     2341 agggcaccca gaccgccaag ctgaaggtga ccaagggcgg ccccctgccc ttcgcctggg
     2401 acatcctgtc ccctcagttc atgtacggct ccaaggccta cgtgaagcac cccgccgaca
     2461 tccccgacta cttgaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact
     2521 tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca
     2581 tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga
     2641 agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg
     2701 gcgagatcaa gcagaggctg aagctgaagg acggcggcca ctacgacgcc gaggtcaaga
     2761 ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc aacatcaagc
     2821 tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgag cgcgccgagg
     2881 gccgccactc caccggcggc atggacgagc tgtacaagta atgatagctc gagagatctc
     2941 ccagctttct tgtacaaagt tggcattata agaaagcatt gcttatcaat ttgttgcaac
     3001 gaacaggtca ctatcagtca aaataaaatc attatttgcc atccagctga tatcccctat
     3061 agtgagtcgt attacatggt catagctgtt tcctggcagc tctggcccgt gtctcaaaat
     3121 ctctgatgtt acattgcaca agataaaaat atatcatcat gaacaataaa actgtctgct
     3181 tacataaaca gtaatacaag gggtgttatg agccatattc aacgggaaac gtcgaggccg
     3241 cgattaaatt ccaacatgga tgctgattta tatgggtata aatgggctcg cgataatgtc
     3301 gggcaatcag gtgcgacaat ctatcgcttg tatgggaagc ccgatgcgcc agagttgttt
     3361 ctgaaacatg gcaaaggtag cgttgccaat gatgttacag atgagatggt cagactaaac
     3421 tggctgacgg aatttatgcc tcttccgacc atcaagcatt ttatccgtac tcctgatgat
     3481 gcatggttac tcaccactgc gatccccgga aaaacagcat tccaggtatt agaagaatat
     3541 cctgattcag gtgaaaatat tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg
     3601 attcctgttt gtaattgtcc ttttaacagc gatcgcgtat ttcgtctcgc tcaggcgcaa
     3661 tcacgaatga ataacggttt ggttgatgcg agtgattttg atgacgagcg taatggctgg
     3721 cctgttgaac aagtctggaa agaaatgcat aaacttttgc cattctcacc ggattcagtc
     3781 gtcactcatg gtgatttctc acttgataac cttatttttg acgaggggaa attaataggt
     3841 tgtattgatg ttggacgagt cggaatcgca gaccgatacc aggatcttgc catcctatgg
     3901 aactgcctcg gtgagttttc tccttcatta cagaaacggc tttttcaaaa atatggtatt
     3961 gataatcctg atatgaataa attgcagttt catttgatgc tcgatgagtt tttctaatca
     4021 gaattggtta attggttgta acactggcag agcattacgc tgacttgacg ggacggcgca
     4081 agctcatgac caaaatccct taacgtgagt tacgcgtcgt tccactgagc gtcagacccc
     4141 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     4201 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     4261 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     4321 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     4381 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     4441 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     4501 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     4561 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     4621 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     4681 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     4741 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     4801 tttgctcaca tgtt
//
