LOCUS       Exported                4631 bp ds-DNA     circular SYN 14-FEB-2018
DEFINITION  synthetic circular DNA
ACCESSION   urn.local...5k-5kkvd3x
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4631)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Feb 14, 2018 from SnapGene 4.1.4
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4631
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     2..21
                     /label=WT019
     modified_base   2..16
                     /label=user junction
                     /note="/created_by=User"
     promoter        131..159
                     /label=amp prom
                     /note="/Imported_from=Pla sMapper"
                     /note="/Transferred_From=amp prom"
                     /note="/Transferred_Similarity=100.00%"
     primer_bind     complement(162..185)
                     /label=WT099 for resistance marker swapping
     primer_bind     171..227
                     /label=WT175
     primer_bind     171..194
                     /label=WT098 for resistance marker swapping
     CDS             201..857
                     /label=cat (CmR) CDS chloramphenicol resistance protein
                     /note="cat (CmR)"
                     /note="from pACYCDuet-1"
                     /note="/modified_by=User"
     primer_bind     complement(758..776)
                     /label=WT181
     primer_bind     765..794
                     /label=WT180
     modified_base   777..778
                     /label=H195A
                     /note="/created_by=User"
     modified_base   785
                     /label=mutation
                     /note="mutation"
                     /note="annotated as a G in pACYCDuet cat marker annotation,
                     here it is an A, but this mutation is silent from codon GTC
                     (Val, 25% codon usage) to GTT (Val, 21% usage), so it 
                     should not be of functional relevance"
     primer_bind     840..857
                     /label=WT083
     primer_bind     complement(842..873)
                     /label=WT176
     modified_base   858..875
                     /label=3GS linker
                     /note="/created_by=User"
     primer_bind     861..896
                     /label=WT177
     misc_feature    876..1688
                     /label=kan2 marker
                     /label=nonstandard type: Marker
                     /note="/Imported_from=Pla sMapper"
     primer_bind     complement(876..895)
                     /label=WT096
     misc_feature    978..980
                     /label=Ser27
                     /note="/created_by=User"
     misc_feature    1074..1076
                     /label=Glu 60
                     /note="/created_by=User"
     misc_feature    1464..1466
                     /label=catalytic Asp
                     /note="/created_by=User"
                     /note="/modified_by=User"
     misc_feature    1479..1481
                     /label=Catalytic Asn
                     /note="/created_by=User"
     misc_feature    1518..1520
                     /label=Catalytic Asp
                     /note="/created_by=User"
     primer_bind     complement(1664..1686)
                     /label=WT100 Kan RP for pWT004c
     primer_bind     1675..1709
                     /label=WT101
     rep_origin      1859..2447
                     /label=pMB1 mutated
                     /note="/created_by=User"
     primer_bind     complement(1859..1879)
                     /label=WT135 seq
     primer_bind     2430..2447
                     /label=WT024 pUC end FP sequencing
     primer_bind     2437..2456
                     /label=PBR322 RP
     primer_bind     complement(2590..2622)
                     /label=WT018
     primer_bind     2608..2628
                     /label=WT017
     modified_base   2608..2622
                     /label=USER junction
                     /note="USER junction"
     terminator      2623..2667
                     /note="rrnB1 transcriptional terminator"
     modified_base   2668..2682
                     /label=USER junction
                     /note="USER junction"
     CDS             complement(2683..3561)
                     /codon_start=1
                     /label=araC
                     /note="/dnas_title=araC"
                     /note="/vntifkey=4"
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     misc_feature    2683..2723
     misc_feature    2734
                     /label=originally an 'a' in annotation
                     /note="originally an 'a' in annotation"
     misc_feature    2773
                     /label=originally a t in annotation
                     /note="originally a t in annotation"
     misc_feature    2839
                     /label=originally an a in annotation
                     /note="originally an a in annotation"
     misc_feature    3043
                     /label=originally a c in annotation
                     /note="originally a c in annotation"
     primer_bind     3116..3133
                     /label=WT046
     misc_feature    3208
                     /label=originally an a in annotation
                     /note="originally an a in annotation"
     misc_feature    3367
                     /label=originally a g in annotation
                     /note="originally a g in annotation"
     misc_feature    3514
                     /label=originally a g in annotation
                     /note="originally a g in annotation"
     primer_bind     3543..3561
                     /label=WT022 seq araC 0 RP
     misc_feature    3577
                     /label=A to G ***
                     /note="/dnas_title=A to G ***"
                     /note="/vntifkey=21"
     protein_bind    3590..3606
                     /note="araO2"
     misc_feature    3659
                     /label=C to A ***
                     /note="/dnas_title=C to A ***"
                     /note="/vntifkey=21"
     prim_transcript 3725
                     /label=pC TSS
                     /note="pC TSS"
     protein_bind    3727..3743
                     /note="araO1"
     protein_bind    3748..3764
                     /note="araO1"
     protein_bind    3769..3790
                     /note="CAP"
     protein_bind    3801..3817
                     /note="araI1"
     protein_bind    3822..3838
                     /note="araI2"
     -35_signal      3835..3840
                     /label=-35
                     /note="-35"
     misc_feature    3835
     misc_feature    3849
     -10_signal      3859..3864
                     /label=-10
                     /note="-10"
     prim_transcript 3873
                     /label=pBAD TSS
                     /note="pBAD TSS"
     primer_bind     complement(3877..3896)
                     /label=WT016
     primer_bind     3882..3930
                     /label=WT020
     modified_base   3882..3896
                     /label=USER junction
                     /note="USER junction"
     RBS             3897..3911
                     /note="SD8 RBS (from Ringquist and Gold Mol. Micro. 1992)"
     CDS             3912..4631
                     /codon_start=1
                     /label=eGFP
                     /note="/created_by=User"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     misc_feature    3912..3916
                     /label=USER junction 1
                     /note="/created_by=User"
     primer_bind     complement(4042..4072)
                     /label=WT182
     sig_peptide     4060..4082
                     /label=eGFP gRNA 1 cut site
                     /note="/created_by=User"
     primer_bind     4062..4080
                     /label=WT183
     misc_feature    complement(4106..4125)
                     /label=does not cut 18h-18a-spacer
                     /note="/created_by=User"
     misc_feature    complement(4111..4130)
                     /label=does not cut 18g-18a-spacer
                     /note="/created_by=User"
                     /note="/modified_by=User"
     misc_feature    4524
                     /label=target C
                     /note="/created_by=User"
     primer_bind     complement(4609..16)
                     /label=WT021
     misc_feature    4624..4631
                     /label=USER junction 2
                     /note="/created_by=User"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atggagaaaa aaatcactgg atataccacc gttgatatat
      241 cccaatggca tcgtaaagaa cattttgagg catttcagtc agttgctcaa tgtacctata
      301 accagaccgt tcagctggat attacggcct ttttaaagac cgtaaagaaa aataagcaca
      361 agttttatcc ggcctttatt cacattcttg cccgcctgat gaatgctcat ccggagttcc
      421 gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct tgttacaccg
      481 ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac gacgatttcc
      541 ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt
      601 tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg gtgagtttca
      661 ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcactatgg
      721 gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatgctg
      781 ccgtttgtga tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg
      841 agtggcaggg cggggcgggc agcggttctg gctccagcca tattcaacgg gaaacgtctt
      901 gctcgaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc
      961 gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag cccgatgcgc
     1021 cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg
     1081 tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta
     1141 ctcctgatga tgcatggtta ctcaccactg cgatccccgg gaaaacagca ttccaggtat
     1201 tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc
     1261 ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg
     1321 ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc
     1381 gtaatggctg gcctgttgaa caagtctgga aagaaatgca taagcttttg ccattctcac
     1441 cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga
     1501 aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg
     1561 ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa
     1621 aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt
     1681 ttttctaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     1741 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     1801 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     1861 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     1921 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     1981 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     2041 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     2101 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     2161 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     2221 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     2281 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     2341 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     2401 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     2461 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     2521 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     2581 gcagccgaac gaccgagcgc agcgagtaag aggacatccg gtcaaataaa acgaaaggct
     2641 cagtcgaaag actgggcctt tcgttttaga cttagggacc ctttatgaca acttgacggc
     2701 tacatcattc actttttctt cacaaccggc acggaactcg ctcgggctgg ccccggtgca
     2761 ttttttaaat acccgcgaga aatagagttg atcgtcaaaa ccaacattgc gaccgacggt
     2821 ggcgataggc atccgggtgg tgctcaaaag cagcttcgcc tggctgatac gttggtcctc
     2881 gcgccagctt aagacgctaa tccctaactg ctggcggaaa agatgtgaca gacgcgacgg
     2941 cgacaagcaa acatgctgtg cgacgctggc gatatcaaaa ttgctgtctg ccaggtgatc
     3001 gctgatgtac tgacaagcct cgcgtacccg attatccatc ggtggatgga gcgactcgtt
     3061 aatcgcttcc atgcgccgca gtaacaattg ctcaagcaga tttatcgcca gcagctccga
     3121 atagcgccct tccccttgcc cggcgttaat gatttgccca aacaggtcgc tgaaatgcgg
     3181 ctggtgcgct tcatccgggc gaaagaaccc cgtattggca aatattgacg gccagttaag
     3241 ccattcatgc cagtaggcgc gcggacgaaa gtaaacccac tggtgatacc attcgcgagc
     3301 ctccggatga cgaccgtagt gatgaatctc tcctggcggg aacagcaaaa tatcacccgg
     3361 tcggcaaaca aattctcgtc cctgattttt caccaccccc tgaccgcgaa tggtgagatt
     3421 gagaatataa cctttcattc ccagcggtcg gtcgataaaa aaatcgagat aaccgttggc
     3481 ctcaatcggc gttaaacccg ccaccagatg ggcattaaac gagtatcccg gcagcagggg
     3541 atcattttgc gcttcagcca tacttttcat actcccacca ttcagagaag aaaccaattg
     3601 tccatattgc atcagacatt gccgtcactg cgtcttttac tggctcttct cgctaaccca
     3661 accggtaacc ccgcttatta aaagcattct gtaacaaagc gggaccaaag ccatgacaaa
     3721 aacgcgtaac aaaagtgtct ataatcacgg cagaaaagtc cacattgatt atttgcacgg
     3781 cgtcacactt tgctatgcca tagcattttt atccataaga ttagcggatc ctacctgacg
     3841 ctttttatcg caactctcta ctgtttctcc atacccgttt tacctgcagg tgcagtaagg
     3901 aggaaaaaaa aatggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     3961 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     4021 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     4081 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     4141 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     4201 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     4261 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     4321 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     4381 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     4441 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     4501 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     4561 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     4621 tgtacaagta a
//
