LOCUS       Exported                6458 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pPDS1-mEGFP-IRES-puro2b
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6458)
  AUTHORS   Held M, Schmitz MH, Fischer B, Walter T, Neumann B, Olma MH, Peter 
            M, Ellenberg J, Gerlich DW
  TITLE     CellCognition: time-resolved phenotype annotation in high-throughput
            live cell imaging.
  JOURNAL   Nat Methods. 2010 Sep . 7(9):747-54.
  PUBMED    20693996
REFERENCE   2  (bases 1 to 6458)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6458
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     regulatory      13..22
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             19..738
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLK
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(64..85)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(325..344)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     intron          808..1037
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     1020..1039
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    1095..1668
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1262..1279)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1489..1508
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             1701..2300
                     /codon_start=1
                     /gene="pac from Streptomyces"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVECPKDRATWCMTRKPGA"
     primer_bind     complement(1701..1720)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2197..2217
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(2436..2453)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2442..2666
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2713..2732
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2792..2808)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2792..2808)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2805..2827)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2816..2832
                     /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)."
     promoter        complement(2840..2870)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2885..2906
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3023..3040)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3194..3782)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3274..3293)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3953..4813)
                     /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     4576..4595
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4814..4918)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4993..5012)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5184..5563
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5564..5767
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5718..5738
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 gatccaccgg tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
       61 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
      121 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
      181 cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc
      241 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
      301 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
      361 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
      421 ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg
      481 gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac
      541 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
      601 ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag
      661 aagcgcgatc acatggtcct taaggagttc gtgaccgccg ccgggatcac tctcggcatg
      721 gacgagctgt acaagtaaag cggccgcata gataactgat ccagtgtgct ggaattaatt
      781 cgctgtctgc gagggccagc tgttggggtg agtactccct ctcaaaagcg ggcatgactt
      841 ctgcgctaag attgtcagtt tccaaaaacg aggaggattt gatattcacc tggcccgcgg
      901 tgatgccttt gagggtggcc gcgtccatct ggtcagaaaa gacaatcttt ttgttgtcaa
      961 gcttgaggtg tggcaggctt gagatctggc catacacttg agtgacaatg acatccactt
     1021 tgcctttctc tccacaggtg tccactccca ggtccaactg caggtcgagc atgcatctag
     1081 ggcggccaat tccgcccctc tccctccccc ccccctaacg ttactggccg aagccgcttg
     1141 gaataaggcc ggtgtgcgtt tgtctatatg tgattttcca ccatattgcc gtcttttggc
     1201 aatgtgaggg cccggaaacc tggccctgtc ttcttgacga gcattcctag gggtctttcc
     1261 cctctcgcca aaggaatgca aggtctgttg aatgtcgtga aggaagcagt tcctctggaa
     1321 gcttcttgaa gacaaacaac gtctgtagcg accctttgca ggcagcggaa ccccccacct
     1381 ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag atacacctgc aaaggcggca
     1441 caaccccagt gccacgttgt gagttggata gttgtggaaa gagtcaaatg gctctcctca
     1501 agcgtattca acaaggggct gaaggatgcc cagaaggtac cccattgtat gggatctgat
     1561 ctggggcctc ggtgcacatg ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc
     1621 cccccgaacc acggggacgt ggttttcctt tgaaaaacac gatgataagc ttgccacaac
     1681 ccacaaggag acgaccttcc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
     1741 acgacgtccc ccgggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
     1801 gccacaccgt cgacccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
     1861 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
     1921 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
     1981 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
     2041 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
     2101 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
     2161 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
     2221 tcggcttcac cgtcaccgcc gacgtcgagt gcccgaagga ccgcgcgacc tggtgcatga
     2281 cccgcaagcc cggtgcctga cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg
     2341 cacgacccca tggctccgac cgaagccgac ccgggcggcc ccgccgaccc cgcacccgcc
     2401 cccgaggccc accgactcta gagctcgctg atcagcctcg actgtgcctt ctagttgcca
     2461 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     2521 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     2581 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     2641 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctcgag
     2701 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     2761 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     2821 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     2881 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     2941 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     3001 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3061 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3121 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3181 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3241 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3301 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     3361 tcccttcggg aagcgtggcg ctttctcaat gctcacgctg taggtatctc agttcggtgt
     3421 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     3481 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     3541 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     3601 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     3661 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     3721 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3781 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3841 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     3901 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     3961 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4021 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4081 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4141 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4201 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4261 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4321 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4381 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     4441 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     4501 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     4561 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     4621 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     4681 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     4741 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     4801 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     4861 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     4921 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
     4981 gtcgactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
     5041 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
     5101 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
     5161 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
     5221 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     5281 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     5341 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggact atttacggta
     5401 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     5461 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     5521 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
     5581 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     5641 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     5701 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     5761 cagagctggt ttagtgaacc gtcagatccg ctagcgctac cggactcaga tctcgagctc
     5821 aagcttcgaa ttctgcagtc gacggtacca tggctactct gatctatgtt gataaggaaa
     5881 atggagaacc aggcacccgt gtggttgcta aggatgggct gaagctgggg tctggacctt
     5941 caatcaaagc cttagatggg agatctcaag tttcaacacc acgttttggc aaaacgttcg
     6001 atgccccacc agccttacct aaagctacta gaaaggcttt gggaactgtc aacagagcta
     6061 cagaaaagtc tgtaaagacc aagggacccc tcaaacaaaa acagccaagc ttttctgcca
     6121 aaaagatgac tgagaagact gttaaagcaa aaagctctgt tcctgcctca gatgatgcct
     6181 atccagaaat agaaaaattc tttcccttca atcctctaga ctttgagagt tttgacctgc
     6241 ctgaagagca ccagattgcg cacctcccct tgagtggagt gcctctcatg atccttgacg
     6301 aggagagaga gcttgaaaag ctgtttcagc tgggcccccc ttcacctgtg aagatgccct
     6361 ctccaccatg ggaatccaat ctgttgcagt ctccttcaag cattctgtcg accctggatg
     6421 ttgaattgcc acctgtttgc tgtgacatag atattcgg
//
