LOCUS       Exported                5137 bp ds-DNA     circular SYN 11-AUG-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pUC19-POLBHR-eGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5137)
  AUTHORS   Robert Sobol
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Aug 23, 2021 from SnapGene 3.3.3
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5137
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    106..127
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding site"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        142..172
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     misc_feature    169..189
                     /note="Sequencing Primer 1"
     protein_bind    180..196
                     /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)."
     primer_bind     204..220
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    218..3640
                     /note="Validated by Sequencing"
     misc_feature    239..1148
                     /note="Upstream Arm"
     misc_feature    239..262
                     /note="Forward Homology Primer- 1427"
     misc_feature    802..822
                     /note="Sequencing Primer 2"
     CDS             1149..1865
                     /codon_start=1
                     /product="enhanced GFP"
                     /note="EGFP"
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     misc_feature    1395..1415
                     /note="Sequencing Primer 3"
     misc_feature    1866..1927
                     /note="POLB Exon 1"
     misc_feature    1887..1889
                     /note="Mutated PAM"
     misc_feature    1890..2734
                     /note="Downstream Arm"
     misc_feature    1998..2018
                     /note="Sequencing Primer 4"
     misc_feature    2164
                     /note="mutated from canonical 'C' to match A549 sequence"
     misc_feature    2586..2607
                     /note="Sequencing Primer 5"
     misc_feature    complement(2713..2734)
                     /note="Reverse Homology Primer- 1428"
     primer_bind     complement(2741..2757)
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3231..3335
                     /gene="bla"
                     /note="AmpR promoter"
     CDS             3336..4196
                     /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"
     rep_origin      4367..4955
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    5136
                     /note="G to C mutation to remove SapI site"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttat
      241 tgcagttcga tatgaaccca ggagttacgc ttgtaaaata aactattgct gatgtgacat
      301 ttccatttct caagcagact gtaaatttct cagatgtagg tcatcatgtt ataattaggg
      361 ttccattgct catgatctag atgaaacact tctccctttc ttgtggtctc ttcattaatt
      421 tggtccctac cgcaaaggca gggggattaa aaataaaata tcttttgaca ctggttggca
      481 ggtgtactaa cacgtgtgcg ggaggattcc tgttctcggc atggttcacg ccgggctccg
      541 gcagcgccct agtacgtaag gggctgcact ggagtgtgga ctgcccggta tccttagaga
      601 aagttttgac agtgtgacgc cagtgtcgcc tctaaccacg tacaaaacgc gccatgcatt
      661 gggtagctat tacagtgcac atttttgtgc aggaagctac cccaccagcg caagtatttc
      721 agaatcaaga tcgcactccc gtttcccctt ctagggaaag gattccagat aaacactgag
      781 aaaacagatt taatatctta ggtcaaaact ccaggtcttc ccataggaag gcccctgggc
      841 tgtcattctg aggtgcctat ttcccccgcg ccgtgtccac gtccacgagt ccacgaaccc
      901 tccgagttct ctctcctcac ctgccgaggg ctcgcccctc tagccccgcc ccgccccgcc
      961 caggacgcgt gacgtcacaa caagcgccgt gcgccccgcc ccccatcggg ggcaaccatt
     1021 gttccgccgg tcgcgccgga gctgggttgc tcctgctccc gtctccaagt cctggtacct
     1081 ccttcaagct gggagagggc tctagtccct ggttctgaac actctggggt tctcgggtgc
     1141 aggccgccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1201 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     1261 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     1321 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     1381 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     1441 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     1501 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1561 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     1621 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     1681 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1741 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     1801 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     1861 acaagatgag caaacggaag gcgccgcaag agactctcaa cgggggaatc accgacatgc
     1921 tcacaggtta gcaccgggcc gggccccgct ggctttcttc tttccttcca gcctcttccc
     1981 ctgccttcct tctctcccac accgacagtc cagtgggtag ggtaggttcc ttgcagcggg
     2041 tcgtcttccg tggggatctc cctccggcgc ccctggctgg ttgtcagtcc tgccggctac
     2101 acctgggcca tcgcttgggc tgcttttggt ctggcccttg gaggaaacgg gtggtcactg
     2161 tcttttagag gctgccatat ccccttccag aaaacagttc tcgtttgttt actcgtggtt
     2221 cttgttcacc cgagccttct gttgcctttc agaactcgca aactttgaga agaacgtgag
     2281 ccaagctatc cacaagtaca atgcttacag gtgggacagt gcagcattct cgggtagcat
     2341 acgttctggg ataccctgtt tagtgtggca attaacagga ctgagggccc agtggatatt
     2401 tggtccatct gcaagagcgg gaaaaagcaa gaatcgaggc tggtacctta ctatttcttg
     2461 aagaatgtgg gcagtgcgtt ataggatcag ttcagataag ttcccaattc tgattgcaga
     2521 tagctgtagc ctgatacgta ttttgtcatc atcaaacatg aattactaga aatgcggtgt
     2581 aattttaacc gtaatgaaag acagcggatg aagtccatac tgtgaaatat atgtggttca
     2641 aggaatcaca ttttttcttt tggttggtct aaagttgatc ctgtgggatc ctgatcctga
     2701 tccttgttcc ttgttcatgg gatcctgtcc ttgtgaattc actggccgtc gttttacaac
     2761 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
     2821 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
     2881 gcctgaatgg cgaatggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     2941 cacaccgcat atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     3001 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     3061 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     3121 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     3181 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     3241 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     3301 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     3361 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     3421 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     3481 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     3541 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     3601 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     3661 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     3721 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     3781 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     3841 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     3901 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     3961 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     4021 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     4081 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     4141 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     4201 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     4261 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     4321 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     4381 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     4441 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     4501 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     4561 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     4621 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     4681 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     4741 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     4801 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     4861 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     4921 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     4981 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     5041 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     5101 ccgagcgcag cgagtcagtg agcgaggaag cggaaca
//
