LOCUS       Exported                5344 bp ds-DNA     circular SYN 04-ABR-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pD090-pUC19 KGA HA5'-mKO2-3'
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5344)
  AUTHORS   Douglas Adamoski
  TITLE     Direct Submission
  JOURNAL   Exported quarta-feira, 4 de abr de 2018 from SnapGene 2.8.3
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5344
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             complement(146..406)
                     /codon_start=1
                     /gene="lacZ"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /note="lacZ-alpha"
                     /translation="SSNSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRP
                     SQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICSDAA"
     primer_bind     379..395
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    396..406
                     /note="MCS"
                     /note="pUC19 multiple cloning site"
     misc_feature    408..1397
                     /note="5' Homology Arm"
     intron          408..1243
     CDS             1244..1397
                     /codon_start=1
                     /gene="GLS"
                     /gene_synonym="AAD20; GAC; GAM; GLS1; KGA"
                     /product="glutaminase kidney isoform, mitochondrial isoform
                     1 precursor"
                     /EC_number="3.5.1.2"
                     /note="GLS"
                     /note="isoform 1 precursor is encoded by transcript variant
                     1; glutaminase C; L-glutamine amidohydrolase; 
                     K-glutaminase; glutaminase, phosphate-activated; 
                     glutaminase kidney isoform, mitochondrial"
                     /db_xref="GI:156104878"
                     /db_xref="CCDS:CCDS2308.1"
                     /db_xref="GeneID:2744"
                     /db_xref="HGNC:HGNC:4331"
                     /db_xref="MIM:138280"
                     /protein_id="NP_055720.3"
                     /translation="WNNTPMDEALHFGHHDVFKILQEYQVQYTPQGDSDNGKENQTVHK
                     NLDGLL"
     misc_signal     1381..1397
                     /note="sgRNA"
     CDS             1407..2060
                     /codon_start=1
                     /note="mKO2"
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     misc_feature    2097..3086
                     /note="3' Homology Arm"
     misc_signal     2097..2099
                     /note="sgRNA"
     CDS             2097..2099
                     /codon_start=1
                     /gene="GLS"
                     /gene_synonym="AAD20; GAC; GAM; GLS1; KGA"
                     /product="glutaminase kidney isoform, mitochondrial isoform
                     1 precursor"
                     /EC_number="3.5.1.2"
                     /note="GLS"
                     /note="isoform 1 precursor is encoded by transcript variant
                     1; glutaminase C; L-glutamine amidohydrolase; 
                     K-glutaminase; glutaminase, phosphate-activated; 
                     glutaminase kidney isoform, mitochondrial"
                     /db_xref="GI:156104878"
                     /db_xref="CCDS:CCDS2308.1"
                     /db_xref="GeneID:2744"
                     /db_xref="HGNC:HGNC:4331"
                     /db_xref="MIM:138280"
                     /protein_id="NP_055720.3"
                     /translation=""
     misc_feature    2100..2102
                     /note="PAM NGG"
     polyA_signal    2277..2282
                     /gene="GLS"
                     /gene_synonym="AAD20; GAC; GAM; GLS1; KGA"
     polyA_site      2310
                     /gene="GLS"
                     /gene_synonym="AAD20; GAC; GAM; GLS1; KGA"
     variation       2590
                     /note="Mutation (A->T)"
     variation       2592
                     /note="Mutation (C->A)"
     variation       2983
                     /note="Mutation (A->G)"
     CDS             complement(3088..3127)
                     /codon_start=1
                     /gene="lacZ"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /note="lacZ-alpha"
                     /translation="MTMITPSLHACRS"
     misc_feature    3088..3110
                     /note="MCS"
                     /note="pUC19 multiple cloning site"
     primer_bind     complement(3123..3139)
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3147..3163
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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(3171..3201)
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     rep_origin      complement(3525..4113)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4284..5144)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5145..5249)
                     /gene="bla"
                     /note="AmpR promoter"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctccag gcttcacagt
      421 ttgtaaagtg taagggaact acccatcgta ccctgtcatt gactagggct gtgagttatg
      481 tagttctgtc tcctcttgca aaagacttac cacttctggc aagtgattaa ccacttctgg
      541 caactcttca tttcttctta tccttgaata ttcatctaca tcactctaaa cagcacagcc
      601 ccagaagcat ggaaagggga gttattagta tggaaagggg agttactctt ctggtgtagt
      661 ggtccgattg agtccatggc ttcccagcct taccagagct gataaaaatg tcaattcctt
      721 tggggccaat cttgctcctc cagtgtgttt tagccctaat gaggtcatgg ttatttctag
      781 acttctgaga cttactgtgg ctttgaattg acacaaacac taattttctg tcaaaggcta
      841 gagtgatgga tgttatatgc ctgaattttc tttgaacatt ttattttaaa agagatactt
      901 tgggcttttc atgaaactaa aattctggaa ccctagagca gtctattata tctgtcacca
      961 caaggacttt ccttaaaaag ctccctccat aagcacttgg atgtgtaaac aagatccttt
     1021 cacagatctt tttctcttcc tctccatatc ccaacttgtc ttggagacta ctttttgaat
     1081 tataaatccc tttctgcatt tctattatta atttttattt gtaaaattgc taaaatagct
     1141 aaatttggcc atttaacctg catttaaaat ctaggaatgt ggggtgatca tctttgtaca
     1201 taaattacct aatgaccctg tccatgctgt gctacgtgtt taggtggaat aacactccca
     1261 tggatgaagc actgcacttt ggacaccatg atgtatttaa aattctccaa gaataccaag
     1321 tccagtacac acctcaagga gattctgaca acgggaagga aaatcaaacc gtccataaga
     1381 atcttgatgg attgttggga tctaccatgg tgagcgtgat caagcccgag atgaagatga
     1441 ggtactacat ggacggctcc gtcaatgggc atgagttcac aatcgagggt gagggcacag
     1501 gcagacctta cgagggacat caggagatga cactgcgcgt cacaatggcc gagggcgggc
     1561 caatgccttt cgccttcgac ctggtgtccc acgtgttctg ttacggccac agagttttta
     1621 ccaagtaccc agaagagatc ccagactatt tcaagcaggc ctttcctgag ggcctgtcct
     1681 gggagaggtc cctggagttc gaggacggcg gctccgcctc cgtgagcgcc cacatcagcc
     1741 tgaggggcaa caccttctac cacaagtcca agttcaccgg cgtgaacttc cccgccgacg
     1801 gccccatcat gcagaaccag agcgtggact gggagccctc caccgagaag atcaccgcca
     1861 gcgacggcgt gctgaagggc gacgtgacca tgtacctgaa gctggagggc ggcggcaacc
     1921 acaagtgcca gatgaagacc acctacaagg ccgccaagga gatcctggag atgcccggcg
     1981 accactacat cggccacagg ctggtgagga agaccgaggg caacatcacc gagcaggtgg
     2041 aggacgccgt ggcccactcc ggtggcggag gctcgggcgg aggtgggtcg ggatcctaat
     2101 ggtctcaaat cccaagattt aaatcactta cctatttaat tgtggaaaat gattatgaag
     2161 aacatgtgta tttctatctg gtagtgatgt atattttaca tttgtcattt cagtgttact
     2221 ggagttttct tcattgtgca cacaggacaa atctgatctc tttgggaaaa aatagaaata
     2281 aaacaatctc cctccataat gtgagcaata ttacctcgtg cattgtataa tttgatgtaa
     2341 aagaaatagt taccaatgct agcttgtgtg gtcttccatg atttatttgt gttttgtgaa
     2401 ttttcaattt atggtgatga tctgctgata tgcatttata aagtaagctc tgttgtacag
     2461 tctgtccaaa tgggtcaagg ttgcctttag aagcaaatag tgtgattttc aagacttcaa
     2521 atacaaattt agtttaagtg tttgaacaac tatatgcact tacggttgtg tgtttaaaat
     2581 gtctctctct caccctagct tcatgatgtg actcttaaaa aactataata gttaacaact
     2641 gttagtaaga tagaccaatt ctgattagac tttatcaggg aatctgttta agatatgttt
     2701 ggtgaccaaa acgtatgtgt gaatgtagtt ataatgcttt tgaaaaattt tcctttttct
     2761 atatcccctt agtccagcct ctcttctcag acatttagct atctgcctct ttcctttagc
     2821 tgggaaagtg agagctggca tactatgcag tttttatgtt ttccatagta agtcagaaaa
     2881 tgcctcctat ttctggcatc agaactttgc catttgtcta cagaagacga accagagaca
     2941 aaattactaa gtataaatta gtcaagttta tcagtctaaa aagcgaaggg atgtgcaact
     3001 gcagctcttt aagaagtttt ttttttttag cttctagggt aaagataaat tcagaaatgc
     3061 tctaagctac caaagttatt ctgaaagtcg acctgcaggc atgcaagctt ggcgtaatca
     3121 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3181 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3241 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3301 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3361 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     3421 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     3481 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     3541 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     3601 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     3661 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     3721 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     3781 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     3841 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     3901 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     3961 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4021 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4081 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4141 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4201 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4261 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4321 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4381 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     4441 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     4501 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     4561 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     4621 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     4681 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     4741 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     4801 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     4861 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     4921 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     4981 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5041 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5101 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5161 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5221 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     5281 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     5341 cgtc
//
