LOCUS       Exported                9275 bp ds-DNA     circular SYN 22-JUL-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9275)
  AUTHORS   The Weissman lab
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Aug 6, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9275
                     /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"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    874..936
                     /label=5'ITR
     misc_feature    complement(1400..1993)
                     /label=WPRE
                     /label=WPRE; Woodchuck post-transcriptional
     primer_bind     complement(1893..1912)
                     /label=HKO083
     CDS             complement(2016..2615)
                     /label=Puro (-)
     promoter        complement(2628..3130)
                     /label=PGK (-)
     misc_feature    3137..6641
                     /label=endogenous COX5B
     misc_feature    5137..5165
                     /label=5'UTR
     misc_feature    5166..5552
                     /label=CDS
     misc_feature    5409..5416
                     /label=mutant sites - for qPCR
     misc_feature    5553..5555
                     /label=stop codon
     misc_feature    5556..5830
                     /label=3'UTR
     misc_feature    6729..6763
                     /label=3'ITR
     primer_bind     complement(7254..7270)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7278..7294
                     /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(7302..7332)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7347..7368
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(7656..8244)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(8415..9275)
                     /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 ataatcatgt gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaac aaaaactcaa
     1081 aatttcttct ataaagtaac aaaactttta tgagggacag ccccccccca aagcccccag
     1141 ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg ggctccgctc
     1201 cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc ccgggcacgg
     1261 ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg agcctgcaga
     1321 cacctggggg gatacgggga aaaggcctcc agggtttatt acagggacag cagagatcca
     1381 gtttggttaa ttagctagcg atgcggggag gcggcccaaa gggagatccg actcgtctga
     1441 gggcgaaggc gaagacgcgg aagaggccgc agagccggca gcaggccgcg ggaaggaagg
     1501 tccgctggat tgagggccga agggacgtag cagaaggacg tcccgcgcag aatccaggtg
     1561 gcaacacagg cgagcagcca aggaaaggac gatgatttcc ccgacaacac cacggaattg
     1621 tcagtgccca acagccgagc ccctgtccag cagcgggcaa ggcaggcggc gatgagttcc
     1681 gccgtggcaa tagggagggg gaaagcgaaa gtcccggaaa ggagctgaca ggtggtggca
     1741 atgccccaac cagtgggggt tgcgtcagca aacacagtgc acaccacgcc acgttgcctg
     1801 acaacgggcc acaactcctc ataaagagac agcaaccagg atttatacaa ggaggagaaa
     1861 atgaaagcca tacgggaagc aatagcatga tacaaaggca ttaaagcagc gtatccacat
     1921 agcgtaaaag gagcaacata gttaagaata ccagtcaatc tttcacaaat tttgtaatcc
     1981 agaggttgat taccgataag cttgatatcg aattcttaac tggcaccggg cttgcgggtc
     2041 atgcaccagg tgcgcggtcc ttcgggcacc tcgacgtcgg cggtgacggt gaagccgagc
     2101 cgctcgtaga aggggaggtt gcggggcgcg gaggtctcca ggaaggcggg caccccggcg
     2161 cgctcggccg cctccactcc ggggagcacg acggcgctgc ccagaccctt gccctggtgg
     2221 tcgggcgaga cgccgacggt ggccaggaac cacgcgggct ccttgggccg gtgcggcgcc
     2281 aggaggcctt ccatctgttg ctgcgcggcc agccgggaac cgctcaactc ggccatgcgc
     2341 gggccgatct cggcgaacac cgcccccgct tcgacgctct ccggcgtggt ccagaccgcc
     2401 acggcggcgc cgtcgtccgc gacccacacc ttgccgatgt cgagcccgac gcgcgtgagg
     2461 aagagttctt gcagctcggt gacccgctcg atgtggcggt ccggatcgac ggtgtggcgc
     2521 gtggcggggt agtcggcgaa cgcggcggcg agggtgcgta cggccctggg gacgtcgtcg
     2581 cgggtggcga ggcgcaccgt gggcttgtac tcggtcatgg tggtaccccc tggggagaga
     2641 ggtcggtgat tcggtcaacg agggagccga ctgccgacgt gcgctccgga ggcttgcaga
     2701 atgcggaaca ccgcgcgggc aggaacaggg cccacactac cgccccacac cccgcctccc
     2761 gcaccgcccc ttcccggccg ctgctctcgg cgcgccctgc tgagcagccg ctattggcca
     2821 cagcccatcg cggtcggcgc gctgccattg ctccctggcg ctgtccgtct gcgagggtac
     2881 tagtgagacg tgcggcttcc gtttgtcacg tccggcacgc cgcgaaccgc aaggaacctt
     2941 cccgacttag gggcggagca ggaagcgtcg ccggggggcc cacaagggta gcggcgaaga
     3001 tccgggtgac gctgcgaacg gacgtgaaga atgtgcgaga cccagggtcg gcgccgctgc
     3061 gtttcccgga accacgccca gagcagccgc gtccctgcgc aaacccaggg ctgccttgga
     3121 aaaggcgcaa acgcgttcca aaggagacct gagggactgg gtgtgagttg acagtggtgg
     3181 gagctggaag cagcaaggga tgcggggaca tctggggagc ctggacctga ccttggagct
     3241 cactcacaca atctaaataa tcccagagtg aggctgaggt tttcctgagt tggatgaggg
     3301 cctgcgaaaa tttcctatta ttaggcaagc ctactatgag agtgagaagc ctaaactcaa
     3361 tttccatttt atagcctgga gtaaaccact tctggtcagg cagcagggaa tcataacata
     3421 tcagagctga atacctcact ggtttgctga ggggcaagtt agtaggtgtt cctcagaaag
     3481 gattttgtgg tcaattaaga cagggaaact ctggcttaaa ctgcaggtaa attaaataag
     3541 aggtctttac tgcaagccta atcagagcct ttattatgct agtgtccatc tccaaaaggg
     3601 tggtcatcaa atgcaggccc tccctgtctt gaccaagtcc tttttcttga gatcatgtct
     3661 tgagacttgg atgccatgca atacactttg ggaaatacta acctagttga acgacagttt
     3721 tactgatgta gaaatggcgg cccaaaggat aaaaagtagc tgaactagct ggtggggcta
     3781 ctttccagac caatgctttt ttctcctgcc taagacctta accacagttg caaattaact
     3841 acagatcctc agaattttcg cccaattcaa cttggagggg ttgcagtgga ggtgagggag
     3901 aagggagtaa cagtgaagcc attcaaagca aaaaccacct ttaaagaaat tttggcctta
     3961 gaatacaggg tttttttttt ttttttttta attggtggag ggggcaccca ggaagaccag
     4021 acgtgctcct gtgaagcaac gctccaccac gtccttttga aaataccgta gcctgcatga
     4081 gacaggcggg agcacctaag ccttcaactc tccccatgtt gcctccaatt gacctcactg
     4141 ggccaggaaa agccctataa actggttttg tatactctat ccaagtagta aatttactaa
     4201 aattactgga tcctactata taccagacct ctgcaaggca ttgaagattc aatgatgagc
     4261 attaattccc actctaaggg gatttttcta tctggtaggg aaactcagat accgcatgca
     4321 gctagcctgt cacagggcta tactgactaa agctcaccct aagaaatttg cacatcttag
     4381 cctatgggca acgggaagcc atggacggaa tatgttttcc gtttcaaaca aggaactcca
     4441 gtcacagcag gctctaactc catttcctat tatcaactcc ccttcctgct cctcctccct
     4501 tagtagcctc gaaaaagagt ggccacttcc ttcacatcac cagggccttg gtcctggtag
     4561 ggggtcccgc ccccggccct gtcctcctcc ccaggccggg acctccgcgc ctcccacaca
     4621 cagtacaact ctaacacgaa actgggccag gactgcaggt aggtcccgtc caatgcgaca
     4681 cgctcgcgac ggcccacgca ctgccagaac cagccctgcc actaaccagc ctagtaagga
     4741 acgcagtgat ctttaggagc ccacttctaa attcaaatca tttggggttc agtaagtctc
     4801 ttcacagacg gggacctaat agcttccatt agcagcctcc acgtttgctg aacacgacta
     4861 cggggccaga cccacaggaa ctccaatgtg gttcagccgc tcaactccag ggaccaaaaa
     4921 gactttccca cctcccagcg ttattaaagg caaccccgcc cccaagcctc agccactacg
     4981 cggtgcagaa agaggcagag ctgggcatgc gcaatttccg cccccgccac agtcttggga
     5041 gtgggcgggc tcgagggctg ggaggaagcc gaaagctgac cgagaggaga aagaagcccg
     5101 cccccggaag tccctcctgt ctctgcagct tgttcccgga agttttgctg ctagtcgcgg
     5161 acgcaatggc ttcaaggtta cttcgcggag ctggaacgct ggccgcgcag gccctgaggg
     5221 ctcgcggccc cagtggcgcg gccgcgatgc gctccatggc atctggaggt ggtgttccca
     5281 ctgatgaaga gcaggcgact gggttggaga gggagatcat gctggctgca aagaagggac
     5341 tggacccata caatgtactg gccccaaagg gagcttcagg caccagggaa gaccctaatt
     5401 tagtccccag catcagcaac aagagaatag taggctgcat ctgtgaagag gacaatacca
     5461 gcgtcgtctg gttttggctg cacaaaggcg aggcccagcg atgcccccgc tgtggagccc
     5521 attacaagct ggtgccccag cagctggcac actgagcacc tgcactaaat tactcaaaat
     5581 gtgctgtaaa gtttcttctt tccagtaaag actagccatt gcattggctc cttctcccat
     5641 agatggctgg tcttatttct tacccgtatt ctttggtagg catggaatat gcttattttg
     5701 ggaaaagctg tctgttaatg ctagcttgcc atccacttac tgaaagtgta taaccagtgt
     5761 atagtgctta gattaataat aagaatagat cgacaacccg taatgcaatg aatgggacca
     5821 cctggtatga gagaaagggg cgggctgagg agtcaggctg acaggactta aaatattggc
     5881 tccatcattt ggctctatct ctgtggacat gttctctggg ggtcaagtac aactacaaaa
     5941 aggtaagtac cttacaaggt gcttaatatg taaagagctg agtggatagt aggtcctcac
     6001 agtatcctca tagagacagg gtggccttgt gaagagagtt ttggggattt tgaagtcttg
     6061 agggacccag gtgcaaagta agaattcatg gggataccag cagcaaaatt cagaatatgg
     6121 aacaatttca gaaatcactg accatgtttc ttcaaataaa ttgcaaggag tggggggaaa
     6181 agctagggag aacatgtgga gaaacagata agggagatac cagtcagcag catgcacacc
     6241 ttgtttagat cttgatttga aaacaattat aaaggaaaaa aaattgagaa gaatttgaac
     6301 tctggatatt tgatatgtag acatttacca ccctttcagg tatttatata gttttgtttt
     6361 gtttttttta atgcttgaga tacgcacagg aataagggag gggagaaatg ccaaaaaagc
     6421 ccactttagc tgcttcatac ctgtgacacc ttgggacaag ttaaaatctc aaaaccctct
     6481 tacctacaaa atggaaatac cttttcattt tgtgtggaga ttagccagag gtggcttcag
     6541 ttactatctt cacaaaattg gtcgaaagta ggtatttcag tgggagctga tagagtcaga
     6601 gaaaagcccc ttcccctcat cttgcccatc ctgaaggtgt agtttaaacc aagcggcgac
     6661 tgagatgtcc taaatgcaca gcgacggatt cgcgctattt agaaagagag agcaatattt
     6721 caagaatgca tgcgtcaatt ttacgcagac tatctttcta gggttaatct agctgcatca
     6781 ggatcatatc gtcgggtctt ttttccggct cagtcatcgc ccaagctggc gctatctggg
     6841 catcggggag gaagaagccc gtgccttttc ccgcgaggtt gaagcggcat ggaaagagtt
     6901 tgccgaggat gactgctgct gcattgacgt tgagcgaaaa cgcacgttta ccatgatgat
     6961 tcgggaaggt gtggccatgc acgcctttaa cggtgaactg ttcgttcagg ccacctggga
     7021 taccagttcg tcgcggcttt tccggacaca gttccggatg gtcagcccga agcgcatcag
     7081 caacccgaac aataccggcg acagccggaa ctgccgtgcc ggtgtgcaga ttaatgacag
     7141 cggtgcggcg ctgggatatt acgtcagcga ggacgggtat cctggctgga tgccgcagaa
     7201 atggacatgg ataccccgtg agttacccgg cgggcgcgct tggcgtaatc atggtcatag
     7261 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     7321 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     7381 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     7441 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     7501 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     7561 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     7621 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     7681 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     7741 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     7801 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     7861 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     7921 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     7981 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     8041 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     8101 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     8161 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     8221 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     8281 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     8341 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     8401 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     8461 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     8521 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     8581 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     8641 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     8701 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     8761 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     8821 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     8881 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     8941 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     9001 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     9061 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     9121 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     9181 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     9241 ggaataaggg cgacacggaa atgttgaata ctcat
//
