LOCUS       Exported                5772 bp ds-DNA     circular SYN 10-AUG-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5772)
  AUTHORS   Jonathan Pacheco
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, May 18, 2023 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5772
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     565..589
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     misc_feature    591
                     /label=MCS
                     /note="multiple cloning site"
     misc_feature    613..1419
                     /label=Tubby(c)
                     /note="Tubby(c)"
     primer_bind     1423..1463
                     /label=FRB-P4C-FWD
     primer_bind     complement(1426..1462)
                     /label=Tubby-FRB-REV
     misc_feature    1431..1441
                     /label=MCS
                     /note="multiple cloning site"
     CDS             1443..1721
                     /codon_start=1
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     primer_bind     complement(1536..1556)
                     /label=FRB rev seq 1
     primer_bind     1619..1639
                     /label=FRB seq fwd 1
     primer_bind     complement(1699..1741)
                     /label=FRB-P4C-REV
     primer_bind     1702..1740
                     /label=Tubby-FRB-FWD
     regulatory      1722..1731
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1728..2438
                     /codon_start=1
                     /product="photoactivatable monomeric derivative of DsRed 
                     fluorescent protein (Subach et al., 2009)"
                     /label=PAmCherry1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHVFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFTWDILSPQFMYGSNAYVKHPADIPDYFKLSFPEGFKWERVMKF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEALSERMYPEDGALK
                     GEVKPRVKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNRKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1731..1748)
                     /label=FPv-R
                     /note="Amplify vector without eGFP/CFP etc pr mCherry etc. 
                     Use to exchange with FPi primers."
     primer_bind     complement(1873..1891)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2141..2160
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2560..2681
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2597..2616)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2651..2670
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2688..3143)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2825..2846)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3037..3056
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3170..3274
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3276..3633
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(3299..3319)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3484..3619
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3546..3565
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(3569..3580)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3668..4462
                     /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(3722..3741)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4332..4351
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4650..4669)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4694..4741
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5070..5658
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5559..5578
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct atggtggata ttgaggtcca ggatctagag gagtttgcac
      661 tgaggccagc cccacaaggg atcaccatca aatgccgcat cactcgggac aagaagggga
      721 tggaccgcgg catgtacccc acctactttc tgcacctaga ccgtgaggat ggcaagaagg
      781 tgttcctcct ggcgggcagg aagagaaaga agagtaaaac ttccaattac ctcatctctg
      841 tggacccaac agacttgtct cggggaggcg atagctatat cgggaaattg cgatccaacc
      901 tgatgggcac caagttcacc gtttatgaca atggcgtcaa ccctcagaag gcatcctctt
      961 ccacgctgga aagcggaacc ttgcgccagg agctggcagc ggtgtgctat gagacaaatg
     1021 tcctaggctt caagggacct cggaagatga gtgtgatcgt cccaggcatg aacatggttc
     1081 atgagagagt ctgtatccgc ccccgcaatg aacatgagac cctgttagca cgctggcaga
     1141 acaagaacac ggagagcatc attgagctgc agaacaagac gccagtctgg aatgatgaca
     1201 cacagtccta tgtacttaac ttccacggcc gtgtcacaca ggcttctgtg aagaacttcc
     1261 agatcatcca cggcaatgac ccggactaca tcgtcatgca gtttggccgg gtagcagaag
     1321 atgtgttcac catggattac aactacccac tgtgtgcact gcaggccttt gccattgctc
     1381 tgtccagctt tgacagcaag ctggcctgcg agacggtacc gcgggcccgg gatccaccgg
     1441 tcatcctctg gcatgagatg tggcatgaag gcctggaaga ggcatctcgt ttgtactttg
     1501 gggaaaggaa cgtgaaaggc atgtttgagg tgctggagcc cttgcatgct atgatggaac
     1561 ggggccccca gactctgaag gaaacatcct ttaatcaggc ctatggtcga gatttaatgg
     1621 aggcccaaga gtggtgcagg aagtacatga aatcagggaa tgtcaaggac ctcacccaag
     1681 cctgggacct ctattatcat gtgttccgac gaatctcaaa ggccaccatg gtgagcaagg
     1741 gcgaggagga taacatggcc atcattaagg agttcatgcg cttcaaggtg cacatggagg
     1801 ggtccgtgaa cggccacgtg ttcgagatcg agggcgaggg cgagggccgc ccctacgagg
     1861 gcacccagac cgccaagctg aaggtgacca agggcggccc cctgcccttc acctgggaca
     1921 tcctgagccc tcagttcatg tacggctcca atgcctacgt gaagcacccc gccgacatcc
     1981 ccgactactt taagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaaattcg
     2041 aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct
     2101 acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtgatg cagaagaaga
     2161 ccatgggctg ggaggccctc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg
     2221 aggtcaagcc cagagtgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca
     2281 cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac cgcaagctgg
     2341 acatcaccag ccacaacgag gactacacca tcgtggagca gtacgagaga gccgagggcc
     2401 gccactccac cggcggcatg gacgagctgt acaagtaagc ggccgcgact ctagatcata
     2461 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     2521 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     2581 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     2641 cattctagtt gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt
     2701 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     2761 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     2821 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     2881 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     2941 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     3001 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     3061 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     3121 taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     3181 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     3241 ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg
     3301 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3361 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3421 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3481 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3541 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     3601 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     3661 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     3721 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     3781 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     3841 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     3901 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3961 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     4021 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     4081 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     4141 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     4201 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     4261 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     4321 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     4381 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     4441 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     4501 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     4561 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     4621 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     4681 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     4741 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     4801 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     4861 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     4921 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     4981 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     5041 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     5101 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     5161 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     5221 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     5281 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     5341 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     5401 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     5461 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     5521 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     5581 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     5641 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     5701 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     5761 accgccatgc at
//
