LOCUS       sequence-340310-        3475 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vitro transcription of PuroR-F2A-mClover3 synthetic construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3475)
  TITLE     Verstreken lab plasmids
REFERENCE   2  (bases 1 to 3475)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pIVTR(T7G)-PuroR-F2A-mClover3
FEATURES             Location/Qualifiers
     source          1..3475
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     373..392
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(691..1404,1405..1407,1408..1410))
                     /codon_start=1
                     /product="bright green-yellow fluorescent protein derived 
                     from mClover2, with improved photostability (Bajar et al., 
                     2003)"
                     /label=mClover3
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(736..757)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1344..1365
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(1411..1476)
                     /codon_start=1
                     /product="2A peptide from foot-and-mouth disease virus 
                     polyprotein"
                     /label=F2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="VKQTLNFDLLKLAGDVESNPGP"
     CDS             complement(1486..2085)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MATEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGSRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1566..1586)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     regulatory      2082..2091
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(2172..2191)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2173..2191)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2223..2327
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2328..2396,2397..3188)
                     /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     complement(2546..2565)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(3359..3475,1..472)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
       61 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
      121 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
      181 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
      241 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
      301 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
      361 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
      421 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
      481 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctccca tggttttgca
      541 atgaaaataa atgtttttta ttaggcagaa tccagatgct caaggccctt cataatatcc
      601 cccagtttag tagttggact tagggaacaa aggaaccttt aatagaaatt ggacagcaag
      661 aaagcgagca agcttgcatg cctgcaggtc ttacttgtac agctcgtcca tgccatgtgt
      721 aatcccggcg gcggtcacga actccagcag gaccatgtga tcgcgcttct cgttggggtc
      781 tttgctcagc ttggactgat ggctcaggta gtggttgtcg ggcagcagca cggggccgtc
      841 gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc acgctgccgt cctcaacgtt
      901 gtggcggatc ttgaagttag ccttgatgca gttcttctgc ttgtcggccg tgatatagac
      961 gtagtggctg ttgaagttgt actccagctt gtgccccagg atgttgccgt cctccttgaa
     1021 gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact tcacctcggc
     1081 gcgggtcttg taggtaccgt cgtccttgaa agagatggtg cgctcctgga cgtagccttc
     1141 gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc ggctgaagca
     1201 ggccacgccg tagccgaagg tggtcacgag ggtgggccag ggcacgggca gcttgccggt
     1261 ggtgcagatg aacttcaggg tcagcttgcc gttggtggca tcgccctcgc cctcgccgcg
     1321 gacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg gcaccacccc
     1381 ggtgaacagc tcctcgccct tagacaccat gggcccaggg ttggactcga cgtctcccgc
     1441 caacttgaga aggtcaaaat tcaaagtctg tttcactcca ctgccggcac cgggcttgcg
     1501 ggtcatgcac caggtgcgcg gtccttcggg cacctcgacg tcggcggtga cggtgaagcc
     1561 gagccgctcg tagaagggga ggttgcgggg cgcggaggtc tccaggaagg cgggcacccc
     1621 agcgcgctcg gccgcctcca ctccggggag cacgacggcg ctgcccagac ccttgccctg
     1681 gtggtcgggc gagacgccga cggtggccag gaaccacgcg ggctccttgg gccggtgcgg
     1741 cgccaggagg ccttccatct gttgctgcgc ggccagccgg gaaccgctca actcggccat
     1801 gcgcgagccg atctcggcga acaccgcccc cgcttcgacg ctctccggcg tggtccagac
     1861 cgccaccgcg gcgccgtcgt ccgcgaccca caccttgccg atgtcgagcc cgacgcgcgt
     1921 gaggaagagt tcttgcagct cggtgacccg ctcgatgtgg cggtccgggt cgacggtgtg
     1981 gcgcgtggcg gggtagtcgg cgaacgcggc ggcgagggtg cgtacggccc gggggacgtc
     2041 gtcgcgggtg gcgaggcgca ccgtgggctt gtactcggtg gccatggtgg cgactctaga
     2101 ggatccccgg gtaccgagct cgaattctgt ttgaggttgc tagtgaacac agttgtgtca
     2161 gaagcaaatg tccctatagt gagtcgtatt agacgtcagg tggcactttt cggggaaatg
     2221 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     2281 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     2341 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     2401 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     2461 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     2521 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     2581 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     2641 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     2701 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     2761 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     2821 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     2881 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     2941 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     3001 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     3061 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     3121 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     3181 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     3241 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     3301 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     3361 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     3421 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actgg
//
