LOCUS       Exported                4814 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    RIP-Timer (DM#285)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4814)
  AUTHORS   Dor Y, Brown J, Martinez OI, Melton DA
  TITLE     Adult pancreatic beta-cells are formed by self-duplication rather 
            than stem-cell differentiation.
  JOURNAL   Nature. 2004 May 6. 429(6987):41-6.
  PUBMED    15129273
REFERENCE   2  (bases 1 to 4814)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4814
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     regulatory      795..804
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             801..1481
                     /codon_start=1
                     /product="green-to-red fluorescent timer derivative of 
                     DsRed (Terskikh et al., 2000)"
                     /label=Timer
                     /note="mammalian codon-optimized"
                     /translation="MVRSSKNVIKEFMRFKVRMEGTVNGHEFEIEGEGEGRPYEGHNTV
                     KLKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     VATVTQDSSLQDGCFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     KALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERTEGRH
                     HLFL"
     primer_bind     complement(984..1004)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     1396..1419
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    1602..1723
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1639..1658)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1693..1712
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1730..2185)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1867..1888)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2079..2098
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2212..2316
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2318..2675
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(2341..2361)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2526..2661
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2588..2607
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2710..3504
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2764..2783)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3374..3393
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3692..3711)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3736..3783
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4112..4700
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4601..4620
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattac tagcgctacc ggactcagat ctcgagctcg gtacccgggg atcccccaac
       61 cactccaagt ggaggctgag aaaggttttg tagctgggta gagtatgtac taagagatgg
      121 agacagctgg ctctgagctc tgaagcaagc acctcttatg gagagttgct gaccttcagg
      181 tgcaaatcta agatactaca ggagaataca ccatggggct tcagcccagt tgactcccga
      241 gtgggctatg ggtttgtgga aggagagata gaagagaagg gacctttctt cttgaattct
      301 gctttccttc tacctctgag ggtgagctgg ggtctcagct gaggtgagga cacagctatc
      361 agtgggaact gtgaaacaac agttcaaggg acaaagttac taggtccccc aacaactgca
      421 gcctcctggg gaatgatgtg gaaaaatgct cagccaagga caaagaaggc ctcaccctct
      481 ctgagacaat gtcccctgct gtgaactggt tcatcaggcc acccaggagc ccctattaag
      541 actctaatta ccctaaggct aagtagaggt gttgttgtcc aatgagcact ttctgcagac
      601 ctagcaccag gcaagtgttt ggaaactgca gcttcagccc ctctggccat ctgctgatcc
      661 acccttaatg ggacaaacag caaagtccag gggtcagggg gggggtgctt tggactataa
      721 agctagtggg gattcagtaa cccccagccc taactctaga gtcgacggta ccgcgggccc
      781 gggatccacc ggtcgccacc atggtgcgct cctccaagaa cgtcatcaag gagttcatgc
      841 gcttcaaggt gcgcatggag ggcaccgtga acggccacga gttcgagatc gagggcgagg
      901 gcgagggccg cccctacgag ggccacaaca ccgtgaagct gaaggtgacc aagggcggcc
      961 ccctgccctt cgcctgggac atcctgtccc cccagttcca gtacggctcc aaggtgtacg
     1021 tgaagcaccc cgccgacatc cccgactaca agaagctgtc cttccccgag ggcttcaagt
     1081 gggagcgcgt gatgaacttc gaggacggcg gcgtggcgac cgtgacccag gactcctccc
     1141 tgcaggacgg ctgcttcatc tacaaggtga agttcatcgg cgtgaacttc ccctccgacg
     1201 gccccgtgat gcagaagaag accatgggct gggaggcctc caccgagcgc ctgtaccccc
     1261 gcgacggcgt gctgaagggc gagatccaca aggccctgaa gctgaaggac ggcggccact
     1321 acctggtgga gttcaagtcc atctacatgg ccaagaagcc cgtgcagctg cccggctact
     1381 actacgtgga caccaagctg gacatcacct cccacaacga ggactacacc atcgtggagc
     1441 agtacgagcg caccgagggc cgccaccacc tgttcctgta gcggccgcga ctctagatca
     1501 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     1561 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     1621 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     1681 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc
     1741 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     1801 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     1861 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     1921 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     1981 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     2041 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     2101 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     2161 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     2221 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     2281 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     2341 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2401 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     2461 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     2521 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     2581 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     2641 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     2701 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     2761 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     2821 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     2881 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     2941 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3001 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3061 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     3121 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     3181 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     3241 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     3301 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     3361 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     3421 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     3481 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     3541 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     3601 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     3661 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     3721 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     3781 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     3841 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     3901 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     3961 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     4021 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4081 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     4141 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     4201 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     4261 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     4321 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     4381 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     4441 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     4501 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     4561 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     4621 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     4681 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     4741 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     4801 ttaccgccat gcat
//
