LOCUS       Exported                9288 bp ds-DNA     circular SYN 23-JUL-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9288)
  AUTHORS   Chen
  TITLE     Direct Submission
  JOURNAL   Exported Jul 24, 2018 from SnapGene Viewer 4.2.1
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9288
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     misc_feature    874..936
                     /label=3' TR
     misc_feature    1113..1344
                     /label=Core Insulator
     misc_feature    1346..1366
                     /label=H1
     misc_feature    1367..1977
                     /label=SH LHA
     misc_feature    1886..1888
                     /label=PAM
     misc_feature    1889..1907
                     /label=SH gRNA 4
     misc_feature    1960..1968
                     /label=SHS231
     misc_feature    1978..2030
                     /label=Bxb1 attP
     promoter        2066..3244
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2297..3235
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     protein_bind    3256..3289
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     CDS             3298..4029
                     /codon_start=1
                     /product="photostable variant of TagRFP (Shaner et al., 
                     2008)"
                     /label=TagRFP-T
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRTD
                     MALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKLNGMDELYK"
     CDS             4036..4089
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4093..5124
                     /codon_start=1
                     /label=HygromycinB Resistance
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     DLQ"
     misc_feature    5132..5173
                     /label=fC31 attP
     misc_feature    5174..5701
                     /label=SH RHA
     misc_feature    5352..5354
                     /label=PAM
     misc_feature    5355..5373
                     /label=SH gRNA 5
     misc_feature    5702..5722
                     /label=H2
     polyA_signal    5863..5980
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    complement(6237..6467)
                     /label=Core Insulator
     misc_feature    6742..6776
                     /label=5' TR
     primer_bind     complement(7267..7283)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7291..7307
                     /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(7315..7345)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7360..7381
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(7669..8257)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(8428..9288)
                     /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"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgc gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaaa caaaaactca
     1081 aaatttcttc tataaagtaa caaaactttt atgagggaca gccccccccc aaagccccca
     1141 gggatgtaat tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct
     1201 ccggtccggc gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg
     1261 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
     1321 acacctgggg ggatacgggg aaaaggcctc caaggccaga ctagaggtac ccattgagtt
     1381 ctgaggtggt ttgtcataca gcaatggcaa tcagatatat ccacacaaat ataattttag
     1441 tttatatttc tgctactgca gttctcatct tattctgagg atatgtgaca aaataattct
     1501 ttcaaaaata ttgatgctgt gctagattac tattttgaat gaattactag acaaatactt
     1561 catatgtatc tttttttttt tttttttttt tttttttttt tttttttgag acggagtctc
     1621 gctctgtcgc ccaggctgga gtgcagtggc gggatctcgg ctcactgcaa tatgtatctt
     1681 attatgtggg tttacacatt atttatctta ttgatttaac ttcaaaacta aactttagtt
     1741 tagctcttgg gccctatctg gaaagggtca tcttttaatc accattaaat cactgaagtc
     1801 atcagtttat tcaaagtact ctgcacaaat tagcattctt tagtggttgt gaaataaata
     1861 gactttaaac ttatcattaa tattcccaat ggtactatgg gggaggcaaa attttctgtc
     1921 ttcttagtgg tttttttttt ttggctaggg ctaaggatta aactgtcaag cttcgaggtg
     1981 gtttgtctgg tcaaccaccg cggtctcagt ggtgtacggt acaaaccccg agtcgacctt
     2041 ctaggtcttg aaaggagtgg gaattggctc cggtgcccgt cagtgggcag agcgcacatc
     2101 gcccacagtc cccgagaagt tggggggagg ggtcggcaat tgaaccggtg cctagagaag
     2161 gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg
     2221 tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt
     2281 tgccgccaga acacaggtaa gtgccgtgtg tggttcccgc gggcctggcc tctttacggg
     2341 ttatggccct tgcgtgcctt gaattacttc cacctggctg cagtacgtga ttcttgatcc
     2401 cgagcttcgg gttggaagtg ggtgggagag ttcgaggcct tgcgcttaag gagccccttc
     2461 gcctcgtgct tgagttgagg cctggcctgg gcgctggggc cgccgcgtgc gaatctggtg
     2521 gcaccttcgc gcctgtctcg ctgctttcga taagtctcta gccatttaaa atttttgatg
     2581 acctgctgcg acgctttttt tctggcaaga tagtcttgta aatgcgggcc aagatctgca
     2641 cactggtatt tcggtttttg gggccgcggg cggcgacggg gcccgtgcgt cccagcgcac
     2701 atgttcggcg aggcggggcc tgcgagcgcg gccaccgaga atcggacggg ggtagtctca
     2761 agctggccgg cctgctctgg tgcctggcct cgcgccgccg tgtatcgccc cgccctgggc
     2821 ggcaaggctg gcccggtcgg caccagttgc gtgagcggaa agatggccgc ttcccggccc
     2881 tgctgcaggg agctcaaaat ggaggacgcg gcgctcggga gagcgggcgg gtgagtcacc
     2941 cacacaaagg aaaagggcct ttccgtcctc agccgtcgct tcatgtgact ccacggagta
     3001 ccgggcgccg tccaggcacc tcgattagtt ctcgagcttt tggagtacgt cgtctttagg
     3061 ttggggggag gggttttatg cgatggagtt tccccacact gagtgggtgg agactgaagt
     3121 taggccagct tggcacttga tgtaattctc cttggaattt gccctttttg agtttggatc
     3181 ttggttcatt ctcaagcctc agacagtggt tcaaagtttt tttcttccat ttcaggtgtc
     3241 gtgaggaatt agcttataac ttcgtatagc atacattata cgaagttatc ctgcaggatg
     3301 gtgtctaagg gcgaagagct gattaaggag aacatgcaca tgaagctgta catggagggc
     3361 accgtgaaca accaccactt caagtgcaca tccgagggcg aaggcaagcc ctacgagggc
     3421 acccagacca tgagaatcaa ggtggtcgag ggcggccctc tccccttcgc cttcgacatc
     3481 ctggctacca gcttcatgta cggcagcaga accttcatca accacaccca gggcatcccc
     3541 gatttcttta agcagtcctt ccctgagggc ttcacatggg agagagtcac cacatacgaa
     3601 gacgggggcg tgctgaccgc tacccaggac accagcctcc aggacggctg cctcatctac
     3661 aacgtcaaga tcagaggggt gaacttccca tccaacggcc ctgtgatgca gaagaaaaca
     3721 ctcggctggg aggccaacac cgagatgctg taccccgctg acggcggcct ggaaggcaga
     3781 accgacatgg ccctgaagct cgtgggcggg ggccacctga tctgcaactt caagaccaca
     3841 tacagatcca agaaacccgc taagaacctc aagatgcccg gcgtctacta tgtggaccac
     3901 agactggaaa gaatcaagga ggccgacaaa gagacctacg tcgagcagca cgaggtggct
     3961 gtggccagat actgcgacct ccctagcaaa ctggggcaca aacttaatgg catggacgag
     4021 ctgtacaagg ctagcgaggg cagaggaagt ctgctaacat gcggtgacgt cgaggagaat
     4081 cctggcccaa ccatgaaaaa gcctgaactc accgctacct ctgtcgagaa gtttctgatc
     4141 gaaaagttcg acagcgtctc cgacctgatg cagctctccg agggcgaaga atctcgggct
     4201 ttcagcttcg atgtgggagg gcgtggatat gtcctgcggg tgaatagctg cgccgatggt
     4261 ttctacaaag atcgctatgt ttatcggcac tttgcatccg ccgctctccc tattcccgaa
     4321 gtgcttgaca ttggggagtt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag
     4381 ggtgtcacct tgcaagacct gcctgaaacc gaactgcccg ctgttctcca gcccgtcgcc
     4441 gaggccatgg atgccatcgc tgccgccgat cttagccaga ccagcgggtt cggcccattc
     4501 ggacctcaag gaatcggtca atacactaca tggcgcgatt tcatctgcgc tattgctgat
     4561 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcctc cgtcgcccag
     4621 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac
     4681 gccgatttcg gctccaacaa tgtcctgacc gacaatggcc gcataacagc cgtcattgac
     4741 tggagcgagg ccatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg
     4801 ccctggttgg cttgtatgga gcagcagacc cgctacttcg agcggaggca tcccgagctt
     4861 gcaggatctc ctcggctccg ggcttatatg ctccgcattg gtcttgacca actctatcag
     4921 agcttggttg acggcaattt cgatgatgca gcttgggctc agggtcgctg cgacgcaatc
     4981 gtccggtccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgctgccgtc
     5041 tggaccgatg gctgtgtgga agtgctcgcc gatagtggaa acagacgccc cagcactcgt
     5101 cctagggcaa aggatctgca gtaatctaga agtgccccaa ctggggtaac ctttgagttc
     5161 tctcagttgg gggactagtg cgtggacaaa gagcagtaaa ttgtgaaaat ggaacaaggc
     5221 aagggggctt agactacagt agttaatcat caagaagtga caaaaaaaat aatggttagt
     5281 taataagatt tgtttaagca gatttctccc agctttagct ctctgtctct ggtgatcaga
     5341 atgcactcct tccttcagac tcagtgagca catcttccac atggaagatt tcttctctag
     5401 cttttaggaa atccagagaa ccctttttgt atctgctgtt tttttttttt taaatgtctt
     5461 taactcaaaa caatttatgt gccaggatga catatctttg gataatgtgt tctgaactcc
     5521 ttcagtacat acgtatataa attaaagcaa atatttttta tgataagctg gcataatagt
     5581 ttcataattt aatcactgat ttaaaaattt aattaaaatt attttttgat attttgtgta
     5641 ataatttttg aggagtatct tttgtgctta atgagtggca gatgacaccc atgttcttag
     5701 ctacgcgtta actaagtaag cggccgcccc ggtatcgaca atcaacctct ggattacaaa
     5761 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     5821 gctgctttaa tgcctttgta tcatgcgtta actaactaaa cttgtttatt gcagcttata
     5881 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     5941 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg aattgactca
     6001 aatgatgtca attagtctat cagaagctat ctggtctccc ttccggggga caagacatcc
     6061 ctgtttaata tttaaacagc agtgttccca aactgggttc ttatatccct tgctctggtc
     6121 aaccaggttg cagggtttcc tgtcctcaca ggaacgaagt ccctaaagaa acagtggcag
     6181 ccaggtttag ccccggaatt gactggattc cttttttagg gcccattggt atggcttttt
     6241 ccccgtatcc ccccaggtgt ctgcaggctc aaagagcagc gagaagcgtt cagaggaaag
     6301 cgatcccgtg ccaccttccc cgtgcccggg ctgtccccgc acgctgccgg ctcggggatg
     6361 cggggggagc gccggaccgg agcggagccc cgggcggctc gctgctgccc cctagcgggg
     6421 gagggacgta attacatccc tgggggcttt gggggggggc tgtccctgat atctataaca
     6481 agaaaatata tatataataa gttatcacgt aagtagaaca tgaaataaca atataattat
     6541 cgtatgagtt aaatcttaaa agtcacgtaa aagataatca tgcgtcattt tgactcacgc
     6601 ggtcgttata gttcaaaatc agtgacactt accgcattga caagcacgcc tcacgggagc
     6661 tccaagcggc gactgagatg tcctaaatgc acagcgacgg attcgcgcta tttagaaaga
     6721 gagagcaata tttcaagaat gcatgcgtca attttacgca gactatcttt ctagggttaa
     6781 tctagctgca tcaggatcat atcgtcgggt cttttttccg gctcagtcat cgcccaagct
     6841 ggcgctatct gggcatcggg gaggaagaag cccgtgcctt ttcccgcgag gttgaagcgg
     6901 catggaaaga gtttgccgag gatgactgct gctgcattga cgttgagcga aaacgcacgt
     6961 ttaccatgat gattcgggaa ggtgtggcca tgcacgcctt taacggtgaa ctgttcgttc
     7021 aggccacctg ggataccagt tcgtcgcggc ttttccggac acagttccgg atggtcagcc
     7081 cgaagcgcat cagcaacccg aacaataccg gcgacagccg gaactgccgt gccggtgtgc
     7141 agattaatga cagcggtgcg gcgctgggat attacgtcag cgaggacggg tatcctggct
     7201 ggatgccgca gaaatggaca tggatacccc gtgagttacc cggcgggcgc gcttggcgta
     7261 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     7321 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     7381 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     7441 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     7501 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     7561 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     7621 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     7681 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     7741 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     7801 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     7861 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     7921 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     7981 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     8041 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     8101 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     8161 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     8221 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     8281 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     8341 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     8401 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     8461 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     8521 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     8581 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     8641 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     8701 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     8761 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     8821 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     8881 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     8941 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     9001 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     9061 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     9121 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     9181 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     9241 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcat
//
