LOCUS       Exported                7528 bp DNA     circular SYN 06-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7528)
  AUTHORS   GICD Gladstone UCSF
  TITLE     Direct Submission
  JOURNAL   Exported Oct 13, 2021 from SnapGene 5.3.1
            https://www.snapgene.com
COMMENT     Colony 4 appears to have a harmless mutation. Colony 3 has a frame 
            shift in the reading frame which is unacceptable. Going with colony 
            5.
FEATURES             Location/Qualifiers
     source          1..7528
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        79..458
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        459..662
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        707..725
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             751..1416
                     /codon_start=1
                     /label=M
                     /translation="MADSNGTITVEELKKLLEQWNLVIGFLFLTWICLLQFAYANRNRF
                     LYIIKLIFLWLLWPVTLACFVLAAVYRINWITGGIAIAMACLVGLMWLSYFIASFRLFA
                     RTRSMWSFNPETNILLNVPLHGTILTRPLLESELVIGAVILRGHLRIAGHHLGRCDIKD
                     LPKEITVATSRTLSYYKLGASQRVAGDSGFAAYSRYRIGNYKLNTDHSSSSDNIALLVQ
                     "
     misc_feature    1428..2001
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     CDS             2039..2265
                     /codon_start=1
                     /label=Envelope
                     /translation="MYSFVSEETGTLIVNSVLLFLAFVVFLLVTLAILTALRLCAYCCN
                     IVNVSLVKPSFYVYSRVKNLNSSRVPDLLV"
     misc_feature    2285..2873
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             complement(2756..2767)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2969..3193
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3239..3667
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        3681..4010
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3861..3996
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4077..4871
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    5045..5166
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5215..5231)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5239..5255
                     /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(5263..5293)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5308..5329
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5617..6205)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6376..7236)
                     /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"
     promoter        complement(7237..7341)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac
       61 gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg
      121 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
      181 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
      241 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
      301 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
      361 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
      421 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
      481 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
      541 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
      601 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
      661 ctctctggct aactagagaa cccactgctt actggcttat cgaaattaat acgactcact
      721 atagggagac ccaagctggc tagcgccacc atggccgact caaatgggac cattactgta
      781 gaggagctga aaaaactgtt ggagcaatgg aaccttgtga ttggatttct cttcttgaca
      841 tggatttgtc tcctgcaatt cgcatacgcg aatcgcaacc gattcctcta tattataaaa
      901 cttatattcc tttggctcct gtggcctgtg acccttgcct gcttcgtact cgcggcggtg
      961 tatcgaataa actggataac tggcggcata gcaatagcta tggcatgcct ggtaggtttg
     1021 atgtggctct cctattttat agcgtcattc agactttttg cacgcacgcg aagcatgtgg
     1081 tcatttaacc ctgaaacaaa tatcttgctt aatgtccccc tgcatggcac aattttgacc
     1141 cgaccgctcc ttgaatcaga attggtgatt ggagccgtga tcctgagggg ccacttgagg
     1201 atcgcaggac atcatcttgg tcgctgtgat attaaggatt tgccaaaaga aataactgta
     1261 gctaccagtc gaactttgag ctattacaag cttggagcgt cccaaagggt cgctggcgat
     1321 agtggatttg cagcctattc acgctacagg atagggaact ataaactgaa tacagaccac
     1381 tcatcatctt ctgataacat tgctttgctc gtccagtaag gatcccgccc ctctccctcc
     1441 ccccccccta acgttactgg ccgaagccgc ttggaataag gccggtgtgc gtttgtctat
     1501 atgttatttt ccaccatatt gccgtctttt ggcaatgtga gggcccggaa acctggccct
     1561 gtcttcttga cgagcattcc taggggtctt tcccctctcg ccaaaggaat gcaaggtctg
     1621 ttgaatgtcg tgaaggaagc agttcctctg gaagcttctt gaagacaaac aacgtctgta
     1681 gcgacccttt gcaggcagcg gaacccccca cctggcgaca ggtgcctctg cggccaaaag
     1741 ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg
     1801 atagttgtgg aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg gctgaaggat
     1861 gcccagaagg taccccattg tatgggatct gatctggggc ctcggtgcac atgctttaca
     1921 tgtgtttagt cgaggttaaa aaaacgtcta ggccccccga accacgggga cgtggttttc
     1981 ctttgaaaaa cacgatgata agcttgccac aacccacaag gagaaattcg ccgccaccat
     2041 gtacagcttc gtatcagaag aaaccgggac actgatcgta aattctgtgc tcttgtttct
     2101 ggcattcgtc gtatttctcc tcgtcacact ggcaattctg actgcattga ggctttgcgc
     2161 ctactgttgt aacattgtca atgtatctct cgtgaaaccc tcattctacg tttacagcag
     2221 ggtgaagaat ctcaattcta gcagggtgcc ggatctcctc gttctttaac tcgaccgact
     2281 cgacaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     2341 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     2401 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     2461 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     2521 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     2581 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     2641 gctgttgggc actgacaatt ccgtggtgtt gtcggggaag ctgacgtcct ttccatggct
     2701 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     2761 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgca
     2821 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcctggaat
     2881 taattcgagc tcggtacctt taagaccaat gacttacaag gcagcctcga gtctagaggg
     2941 cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     3001 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     3061 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     3121 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     3181 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     3241 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     3301 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     3361 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3421 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     3481 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3541 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     3601 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3661 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     3721 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     3781 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     3841 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     3901 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
     3961 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
     4021 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
     4081 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     4141 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     4201 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     4261 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     4321 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     4381 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     4441 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4501 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     4561 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     4621 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4681 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     4741 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     4801 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     4861 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     4921 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4981 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     5041 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     5101 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5161 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     5221 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5281 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5341 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5401 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5461 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5521 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5581 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5641 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5701 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5761 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5821 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5881 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5941 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     6001 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     6061 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6121 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6181 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6241 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6301 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6361 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6421 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6481 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6541 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6601 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6661 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6721 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6781 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6841 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6901 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6961 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7021 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7081 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7141 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7201 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7261 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7321 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     7381 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
     7441 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
     7501 atttaagcta caacaaggca aggcttga
//
