LOCUS       Exported                6072 bp ds-DNA     circular SYN 07-APR-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6072)
  AUTHORS   Wythe Lab
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Jul 22, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6072
                     /organism="synthetic DNA construct"
                     /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=Escherichia coli rrnB terminator
                     /note="rrnB T1 terminator"
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     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"
     misc_feature    691..696
                     /label=mini Kozak
     CDS             697..942
                     /codon_start=1
                     /label=hs_GalT
                     /note="hs_GalT"
                     /translation="MRLREPLLSGSAAMPGASLQRACRLLVAVCALHLGVTLVYYLAGR
                     DLSRLPQLVGVSTPLQGGSNSAAAIGQSSGELRTGGA"
     CDS             970..1668
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     CDS             970..1668
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     primer_bind     995..1018
                     /label=oJDW 1223
     primer_bind     complement(995..1018)
                     /label=TagBFP REV 3
     primer_bind     complement(1082..1103)
                     /label=oJDW 1226 (REV-mTagBFP2-2)
     primer_bind     1584..1606
                     /label=oJDW 1224 / 1672
     primer_bind     complement(1646..1668)
                     /label=oJDW 1225 (REV-mTagBFP2-1)
     CDS             1669..1698
                     /codon_start=1
                     /label=HA
                     /note="HA"
                     /translation="YDVPDYASLP"
     CDS             1714..1770
                     /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     complement(1737..1755)
                     /label=oJDW 1715 (P2A REV2)
     CDS             1777..2160
                     /codon_start=1
                     /label=H2A.F/Z
                     /note="H2A.F/Z"
                     /translation="MAGGKAGKDSGKAKTKAVSRSQRAGLQFPVGRIHRHLKTRTTSHG
                     RVGATAAVYSAAILEYLTAEVLELAGNASKDLKVKRITPRHLQLAIRGDEELDSLIKAT
                     IAGGGVIPHIHKSLIGKKGQQKTA"
     misc_feature    1836
                     /label=silent mutation to destroy BglII site
                     /note="silent mutation to destroy BglII site"
     primer_bind     1922..1943
                     /label=oJDW 1583 (H2A FWD)
     primer_bind     2108..2135
                     /label=JDW (H2A c-term FWDSEQ2)
     primer_bind     complement(2110..2136)
                     /label=oJDW 1582 (H2A REV)
     primer_bind     2121..2146
                     /label=oJDW 1723 (H2A/FZ 3' FWD)
     CDS             2179..4152
                     /codon_start=1
                     /label=tdiRFP
                     /note="tdiRFP"
                     /translation="MAEGSVARQPDLLTCDDEPIHIPGAIQPHGLLLALAADMTIVAGS
                     DNLPELTGLAIGALIGRSAADVFDSETHNRLTIALAEPGAAVGAPITVGFTMRKDAGFI
                     GSWHRHDQLIFLELEPPQRDVAEPQAFFRRTNSAIRRLQAAETLESACAAAAQEVRKIT
                     GFDRVMIYRFASDFSGEVIAEDRCAEVESKLGLHYPASTVPAQARRLYTINPVRIIPDI
                     NYRPVPVTPDLNPVTGRPIDLSFAILRSVSPVHLEFMRNIGMHGTMSISILRGERLWGL
                     IVCHHRTPYYVDLDGRQACELVAQVLAWQIGVMKEGHGTGSTGSGSSGRSGTASSEDNN
                     MAAEGSVARQPDLLTCDDEPIHIPGAIQPHGLLLALAADMTIVAGSDNLPELTGLAIGA
                     LIGRSAADVFDSETHNRLTIALAEPGAAVGAPITVGFTMRKDAGFIGSWHRHDQLIFLE
                     LEPPQRDVAEPQAFFRRTNSAIRRLQAAETLESACAAAAQEVRKITGFDRVMIYRFASD
                     FSGEVIAEDRCAEVESKLGLHYPASTVPAQARRLYTINPVRIIPDINYRPVPVTPDLNP
                     VTGRPIDLSFAILRSVSPVHLEFMRNIGMHGTMSISILRGERLWGLIVCHHRTPYYVDL
                     DGRQACELVAQVLAWQIGVMKEE"
     misc_feature    2179..3126
                     /label=iRFP
                     /note="iRFP"
     primer_bind     2179..2197
                     /label=oJDW 1639 (tdiRFP_FWD1)
     primer_bind     2182..2197
                     /label=Primer 1
     primer_bind     complement(2337..2358)
                     /label=oJDW 1716 (tdiRFP_REV2)
     primer_bind     2792..2818
                     /label=oJDW 1640 (tdIRFP_FWD2)
     misc_feature    2901
                     /label=C>T to destroy ClaI site
     primer_bind     complement(2965..2987)
                     /label=oJDW 1617 (tdiRFP_REV1)
     primer_bind     complement(3104..3127)
                     /label=Primer 2
     primer_bind     3118..3138
                     /label=oJDW 1743 (tdiRFP_FWD3)
     misc_feature    3127..3204
                     /label=Linker
                     /note="Linker"
     primer_bind     complement(3140..3160)
                     /label=oJDW 1835 (tdIRFP linker REVSEQ)
     primer_bind     3161..3220
                     /label=Primer 1
     misc_feature    3205..4149
                     /label=iRFP
                     /note="iRFP"
     primer_bind     3205..3220
                     /label=oJDW 1639 (tdiRFP_FWD1)
     primer_bind     complement(3360..3381)
                     /label=oJDW 1716 (tdiRFP_REV2)
     primer_bind     3815..3841
                     /label=oJDW 1640 (tdIRFP_FWD2)
     misc_feature    3924
                     /label=C>T to destroy ClaI site
     primer_bind     complement(3988..4010)
                     /label=oJDW 1617 (tdiRFP_REV1)
     primer_bind     complement(4127..4161)
                     /label=Primer 2
     protein_bind    complement(4197..4296)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(4314..4332)
                     /label=T7 promoter
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4337..4353)
                     /label=M13 rev
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4466..5275
                     /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      5422..6010
                     /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 caacacattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctcc ttaattaaga attctctaga gccaccatga ggcttcggga gccgctcctg
      721 agcggcagcg ccgcgatgcc aggcgcgtcc ctacagcggg cctgccgcct gctcgtggcc
      781 gtctgcgctc tgcaccttgg cgtcaccctc gtttactacc tggctggccg cgacctgagc
      841 cgcctgcccc aactggtcgg agtctccaca ccgctgcagg gcggctcgaa cagtgccgcc
      901 gccatcgggc agtcctccgg ggagctccgg accggagggg ccaaggatcc acccgtcgcc
      961 accatcgata tgagcgagct gattaaggag aacatgcaca tgaagctgta catggagggc
     1021 accgtggaca accatcactt caagtgcaca tccgagggcg aaggcaagcc ctacgagggc
     1081 acccagacca tgagaatcaa ggtggtcgag ggcggccctc tccccttcgc cttcgacatc
     1141 ctggctacta gcttcctcta cggcagcaag accttcatca accacaccca gggcatcccc
     1201 gacttcttca agcagtcctt ccctgagggc ttcacatggg agagagtcac cacatacgaa
     1261 gacgggggcg tgctgaccgc tacccaggac accagcctcc aggacggctg cctcatctac
     1321 aacgtcaaga tcagaggggt gaacttcaca tccaacggcc ctgtgatgca gaagaaaaca
     1381 ctcggctggg aggccttcac cgagacgctg taccccgctg acggcggcct ggaaggcaga
     1441 aacgacatgg ccctgaagct cgtgggcggg agccatctga tcgcaaacgc caagaccaca
     1501 tatagatcca agaaacccgc taagaacctc aagatgcctg gcgtctacta tgtggactac
     1561 agactggaaa gaatcaagga ggccaacaac gagacctacg tcgagcagca cgaggtggca
     1621 gtggccagat actgcgacct ccctagcaaa ctggggcaca agcttaatta tgacgtccca
     1681 gattacgcaa gtttgcccag atctggcagc ggcgccacga acttctctct gttaaagcaa
     1741 gcaggcgacg tggaagaaaa ccccggtccc ggtaccatgg caggtggaaa agcaggtaaa
     1801 gacagtggaa aagccaagac gaaagccgtc tccaggtctc agagagctgg tctgcagttc
     1861 cctgtaggtc gtatccacag gcacctgaag acccgaacca ccagccatgg ccgtgtcgga
     1921 gctacagctg cggtttacag tgctgctatc ctggagtatc tcaccgctga ggtgctggag
     1981 ttggctggaa acgcttctaa ggatctgaag gtgaagcgaa tcactcccag acatttacag
     2041 ctggccatcc gcggagacga ggagctggac tcgcttatca aggccaccat tgctggagga
     2101 ggagtcattc ctcacatcca caagtctctg attggtaaga agggccagca gaaaaccgca
     2161 gatccaccgg tcgacaccat ggccgaaggg agcgtggcaa gacagcccga tctgctgaca
     2221 tgcgacgacg aaccaatcca cattccagga gctatccagc ctcatggact gctgctggct
     2281 ctggccgctg acatgactat cgtggcaggc tctgataacc tgccagagct gaccgggctg
     2341 gctatcggag cactgattgg ccggtccgca gccgacgtgt tcgattctga gactcacaat
     2401 agactgacca tcgccctggc tgaacctggg gctgcagtgg gagccccaat taccgtcggc
     2461 ttcacaatgc gaaaggacgc tggctttatc gggtcctggc accgccatga tcagctgatt
     2521 tttctggagc tggaaccacc tcagagagac gtggcagagc cacaggcttt ctttcggaga
     2581 acaaacagcg ccatcaggcg cctgcaggcc gctgagactc tggaatccgc ttgcgcagcc
     2641 gctgcacagg aagtgaggaa gatcaccggg ttcgatcgcg tcatgatcta cagattcgcc
     2701 tcagacttta gcggagaagt gatcgcagaa gatcgctgtg ccgaggtcga aagtaaactg
     2761 ggcctgcatt acccagcctc aaccgtgcct gcacaggcac gacggctgta tacaatcaac
     2821 cccgtgcgga tcattcccga cattaattac agaccagtgc ccgtcacacc tgacctgaac
     2881 ccagtgactg ggaggcccat tgatctgagc ttcgccattc tgcgcagcgt gtcccctgtc
     2941 cacctggagt ttatgaggaa tatcggaatg catggcacca tgtctatcag tattctgaga
     3001 ggcgaaaggc tgtgggggct gatcgtgtgc caccatcgga caccttacta tgtggacctg
     3061 gacggcaggc aggcttgtga actggtggct caggtgctgg cttggcagat tggcgtgatg
     3121 aaggaggggc atggcaccgg cagcaccggc agcggcagca gcggccgctc cggcaccgcc
     3181 tcctccgagg acaacaacat ggccgccgaa gggagcgtgg caagacagcc cgatctgctg
     3241 acatgcgacg acgaaccaat ccacattcca ggagctatcc agcctcatgg actgctgctg
     3301 gctctggccg ctgacatgac tatcgtggca ggctctgata acctgccaga gctgaccggg
     3361 ctggctatcg gagcactgat tggccggtcc gcagccgacg tgttcgattc tgagactcac
     3421 aatagactga ccatcgccct ggctgaacct ggggctgcag tgggagcccc aattaccgtc
     3481 ggcttcacaa tgcgaaagga cgctggcttt atcgggtcct ggcaccgcca tgatcagctg
     3541 atttttctgg agctggaacc acctcagaga gacgtggcag agccacaggc tttctttcgg
     3601 agaacaaaca gcgccatcag gcgcctgcag gccgctgaga ctctggaatc cgcttgcgca
     3661 gccgctgcac aggaagtgag gaagatcacc gggttcgatc gcgtcatgat ctacagattc
     3721 gcctcagact ttagcggaga agtgatcgca gaagatcgct gtgccgaggt cgaaagtaaa
     3781 ctgggcctgc attacccagc ctcaaccgtg cctgcacagg cacgacggct gtatacaatc
     3841 aaccccgtgc ggatcattcc cgacattaat tacagaccag tgcccgtcac acctgacctg
     3901 aacccagtga ctgggaggcc cattgatctg agcttcgcca ttctgcgcag cgtgtcccct
     3961 gtccacctgg agtttatgag gaatatcgga atgcatggca ccatgtctat cagtattctg
     4021 agaggcgaaa ggctgtgggg gctgatcgtg tgccaccatc ggacacctta ctatgtggac
     4081 ctggacggca ggcaggcttg tgaactggtg gctcaggtgc tggcttggca gattggcgtg
     4141 atgaaggagg aataactcga gtaatgataa actagtgttt aaactacgta actaggaccc
     4201 agctttcttg tacaaagttg gcattataag aaagcattgc ttatcaattt gttgcaacga
     4261 acaggtcact atcagtcaaa ataaaatcat tatttgccat ccagctgata tcccctatag
     4321 tgagtcgtat tacatggtca tagctgtttc ctggcagctc tggcccgtgt ctcaaaatct
     4381 ctgatgttac attgcacaag ataaaataat atcatcatga acaataaaac tgtctgctta
     4441 cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cgaggccgcg
     4501 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
     4561 gcaatcaggt gcgacaatct atcgcttgta tgggaagccc gatgcgccag agttgtttct
     4621 gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca gactaaactg
     4681 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
     4741 atggttactc accactgcga tccccggaaa aacagcattc caggtattag aagaatatcc
     4801 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
     4861 tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc
     4921 acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta atggctggcc
     4981 tgttgaacaa gtctggaaag aaatgcataa acttttgcca ttctcaccgg attcagtcgt
     5041 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
     5101 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
     5161 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
     5221 taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt tctaatcaga
     5281 attggttaat tggttgtaac actggcagag cattacgctg acttgacggg acggcgcaag
     5341 ctcatgacca aaatccctta acgtgagtta cgcgtcgttc cactgagcgt cagaccccgt
     5401 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     5461 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5521 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     5581 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     5641 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     5701 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     5761 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     5821 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     5881 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     5941 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     6001 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     6061 tgctcacatg tt
//
