LOCUS       pTWIST_CMV_Puro         8165 bp    DNA     circular SYN 11-JUL-2025
DEFINITION  1150.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8165)
  AUTHORS   Alan Monziani
  TITLE     Direct Submission
  JOURNAL   Exported Jul 11, 2025 from SnapGene 8.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    66..187
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     terminator      302..348
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
                     /note="color: #ffffff"
     terminator      378..514
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
                     /note="color: #ffffff"
     misc_feature    559..630
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
                     /note="color: #a6acb3"
     rep_origin      complement(856..1444)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     promoter        1944..3153
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
                     /note="color: #ffffff; direction: RIGHT"
     CDS             3157..3756
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /note="color: #ccffcc"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     polyA_signal    3783..3831
                     /note="synthetic polyadenylation signal"
                     /note="color: #a6acb3"
     CDS             join(4097..4165,4166..4957)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 4097 .. 4165 / #ccffcc / signal sequence
                        2: 4166 .. 4957 / #ccffcc
                     Cleavage site after base 4165"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     terminator      4970..5106
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
                     /note="color: #ffffff"
     misc_feature    5258..5329
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
                     /note="color: #a6acb3"
     terminator      5334..5380
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
                     /note="color: #ffffff"
     polyA_signal    5398..5446
                     /note="synthetic polyadenylation signal"
                     /note="color: #a6acb3"
     enhancer        5527..5830
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        5832..6034
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     gene            6105..8165
                     /label=EPB41L4A ORF
                     /note="color: #99cc00; direction: RIGHT"
ORIGIN
        1 ggatccgcag gcctctgcta gcttgactga ctgagataca gcgtaccttc agctcacaga
       61 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
      121 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
      181 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga
      241 ggttttttaa agcaagtaaa acctctacaa atgtggtatt ggcccatctc tatcggtatc
      301 gtagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgtg cccctcgggc
      361 cggattgcta tctaccggca ttggcgcaga aaaaaatgcc tgatgcgacg ctgcgcgtct
      421 tatactccca catatgccag attcagcaac ggatacggct tccccaactt gcccacttcc
      481 atacgtgtcc tccttaccag aaatttatcc ttaaggtcgt cagctatcct gcaggcgatc
      541 tctcgatttc gatcaagaca ttcctttaat ggtcttttct ggacaccact aggggtcaga
      601 agtagttcat caaactttct tccctcccta atctcattgg ttaccttggg ctatcgaaac
      661 ttaattaacc agtcaagtca gctacttggc gagatcgact tgtctgggtt tcgactacgc
      721 tcagaattgc gtcagtcaag ttcgatctgg tccttgctat tgcacccgtt ctccgattac
      781 gagtttcatt taaatcatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
      841 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
      901 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
      961 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     1021 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     1081 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     1141 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     1201 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     1261 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     1321 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     1381 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     1441 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     1501 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     1561 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     1621 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     1681 atttaaattt ccgaactctc caaggccctc gtcggaaaat cttcaaacct ttcgtccgat
     1741 ccatcttgca ggctacctct cgaacgaact atcgcaagtc tcttggccgg ccttgcgcct
     1801 tggctattgc ttggcagcgc ctatcgccag gtattactcc aatcccgaat atccgagatc
     1861 gggatcaccc gagagaagtt caacctacat cctcaatccc gatctatccg agatccgagg
     1921 aatatcgaaa tcggggcgcg cctggcctcc gcgccgggtt ttggcgcctc ccgcgggcgc
     1981 ccccctcgtc acggcgagcg ctgccacgtc agacgaaggg cgcaggagcg tcctgatcct
     2041 tccgcccgga cgctcaggac agcggcccgc tgctcataag actcggcctt agaaccccag
     2101 tatcagcaga aggacatttt aggacgggac ttgggtgact ctagggcact ggttttcttt
     2161 ccagagagcg gaacaggcga ggaaaagtag tcccttctcg gcgattctgc ggagggatct
     2221 ccgtggggcg gtgaacgccg atgattatat aaggacgcgc cgggtgtggc acagctagtt
     2281 ccgtcgcagc cgggatttgg gtcgcggttc ttgtttgtgg atcgctgtga tcgtcacttg
     2341 gtgagtagcg ggctgctggg ctggccgggg ctttcgtggc cgccgggccg ctcggtggga
     2401 cggaagcgtg tggagagacc gccaagggct gtagtctggg tccgcgagca aggttgccct
     2461 gaactggggg ttggggggag cgcagcaaaa tggcggctgt tcccgagtct tgaatggaag
     2521 acgcttgtga ggcgggctgt gaggtcgttg aaacaaggtg gggggcatgg tgggcggcaa
     2581 gaacccaagg tcttgagccc ttcgctaatg cgggaaagct cttattcggg tgagatgggc
     2641 tgggcaccat ctggggaccc tgacgtgaag tttgtcactg actggagaac tcggtttgtc
     2701 gtctgttgcg ggggcggcag ttatggcggt gccgttgggc agtgcacccg tacctttggg
     2761 agcgcgcgcc ctcgtcgtgt cgtgacgtca cccgttctgt tggcttataa tgcagggtgg
     2821 ggccacctgc cggtaggtgt gcggtaggct tttctccgtc gcaggacgca gggttcgggc
     2881 ctagggtagg ctctcctgaa tcgacaggcg ccggacctct ggtgagggga gggataagtg
     2941 aggcgtcagt ttctttggtc ggttttatgt acctatcttc ttaagtagct gaagctccgg
     3001 ttttgaacta tgcgctcggg gttggcgagt gtgttttgtg aagtttttta ggcacctttt
     3061 gaaatgtaat catttgggtc aatatgtaat tttcagtgtt agacttgtaa attgtccgct
     3121 aaattctggc cgtttttggc ttttttgtta gacaacatga ccgagtacaa gcccacggtg
     3181 cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc
     3241 gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag
     3301 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     3361 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     3421 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     3481 atggagggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     3541 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     3601 gaggccgccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     3661 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     3721 cgcacctggt gcatgacccg caagcccggt gcctaggcta gtatgtaagc ctagtcttag
     3781 ataataaaat cgctatccat cgaagatgga tgtgtgttgg ttttttgtgt gtgtaacgct
     3841 aggcgcgcct ggtgtaccga gaacgatcct ctcagtgcga gtctcgacga tccatatcgt
     3901 tgcttggcag tcagccagtc ggaatccagc ttgggaccca ggaagtccaa tcgtcagata
     3961 ttgtactcaa gcctggtcac ggcagcgtac cgatctgttt aaacctagat attgatagtc
     4021 tgatcggtca acgtataatc gagtcctagc ttttgcaaac atctatcaag agacaggatc
     4081 agcaggaggc tttcgcatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     4141 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     4201 agatcagttg ggtgcgcgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     4261 tgagagtttt cgccccgaag aacgctttcc aatgatgagc acttttaaag ttctgctatg
     4321 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     4381 ttctcagaat gacttggttg agtattcacc agtcacagaa aagcatctta cggatggcat
     4441 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     4501 acttctgaca acgattggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     4561 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     4621 gcgtgacacc acgatgcctg tagcaatggc aacaaccttg cgtaaactat taactggcga
     4681 actacttact ctagcttccc ggcaacagtt gatagactgg atggaggcgg ataaagttgc
     4741 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     4801 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     4861 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     4921 cgctgagata ggtgcctcac tgattaagca ttggtaaccg attctaggtg cattggcgca
     4981 gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc cacatatgcc agattcagca
     5041 acggatacgg cttccccaac ttgcccactt ccatacgtgt cctccttacc agaaatttat
     5101 ccttaagatc ccgaatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga
     5161 gcttacgcga acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc
     5221 gtcagcttac ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca
     5281 ggtatcttct tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa
     5341 cccctcaaga cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac
     5401 acaaaaaacc aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat
     5461 cgagtgtagc cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga
     5521 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
     5581 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
     5641 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
     5701 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
     5761 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
     5821 tattaccatg ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
     5881 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
     5941 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
     6001 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc
     6061 tttgtcgatc ctaccatcca ctcgacacac ccgccagcgg ccgcatgggc tgtttctgcg
     6121 ctgttccgga agaattttac tgcgaagttt tgctcctgga tgaatccaag ttaaccctta
     6181 ccacccagca gcagggcatc aagaagtcaa cgaaaggttc cgttgtcctt gaccacgtat
     6241 tccatcacgt aaaccttgtg gagatagatt attttgggct acgttactgt gacagaagcc
     6301 atcagacgta ttggctggat cctgcaaaaa cccttgctga acacaaagaa ctgatcaaca
     6361 ctggacctcc atatactttg tattttggta ttaaattcta tgctgaagat ccatgtaaac
     6421 ttaaagaaga aataaccaga tatcagtttt tcttgcaggt gaagcaagat gtccttcagg
     6481 gccgtctgcc ctgtcccgtc aacactgctg ctcagctggg agcgtatgcc atccagtcgg
     6541 agcttggaga ttatgaccca tataaacata ctgcaggata tgtatctgag taccggtttg
     6601 ttcctgatca gaaggaagaa cttgaagaag ccatagaaag gattcataaa actctaatgg
     6661 gtcagattcc ttctgaggct gagctgaatt acttgaggac tgccaaatcc ctggagatgt
     6721 atggcgttga cctccatccc gtctatggag aaaacaagtc tgagtatttc ttaggattaa
     6781 ctccggttgg tgttgttgtg tacaagaata aaaagcaagt ggggaagtat ttctggcctc
     6841 ggattacaaa ggttcacttc aaggagactc aatttgaact cagagtactg ggaaaagatt
     6901 gtaacgaaac ctcattcttt tttgaagctc ggagtaaaac tgcttgcaag cacctctgga
     6961 agtgcagtgt ggaacatcat acatttttta gaatgccaga aaatgaatcc aattcactgt
     7021 caagaaaact cagcaagttt ggatccatac gttataagca ccgctacagt ggcaggacag
     7081 ctttgcaaat gagccgagat ctttctattc agcttccccg gcctgatcag aatgtgacaa
     7141 gaagtcgaag caagacttac cctaagcgaa tagcacaaac acagccagct gaatcaaaca
     7201 gcatcagtag gataactgca aacatggaaa atggagaaaa tgaaggaaca attaaaatta
     7261 ttgcaccttc accagtaaaa agctttaaga aagcaaagaa tgaaaatagc cctgataccc
     7321 aaagaagcaa atctcatgca ccgtgggaag aaaatggccc ccagagtgga ctctacaatt
     7381 ctcccagtga tcgcactaag tcgccaaagt tcccttacac gcgtcgccga aacccctcct
     7441 gtggaagtga caatgattct gtacagcctg tgaggaggag gaaagcccat aacagtggtg
     7501 aagattcaga tcttaagcaa aggaggaggt cacgttcacg ctgtaacacc agcagtggta
     7561 gtgaatcaga aaattctaat agagaatacc ggaaaaagag aaacagaata cggcaggaga
     7621 atgatatggt tgattcagcg cctcagtggg aagctgtatt aaggagacaa aaggaaaaaa
     7681 accaagccga ccccaacaac aggcgatcca gacacagatc tcgttcgaga agccccgata
     7741 tccaagcaaa agaagagtta tggaagcaca ttcaaaaaga acttgtggat ccatccggat
     7801 tgtccgaaga acaattaaaa gagattccat acactaaaat agagacacaa ggtgacccaa
     7861 tccgcatcag gcattctcat tcgccacgaa gttaccgcca gtatcgcagg tcccagtgtt
     7921 cagatgggga gcgatcagtt ctctcggaag tgaattcaaa aacagatctt gtaccaccac
     7981 ttccggtgac ccgttcttcg gatgctcagg gttctgggga tgctacagtt catcagagaa
     8041 gaaatgggtc taaagatagc ctgatggaag aaaaacctca gacatctaca aacaacctgg
     8101 ctggaaaaca cacagcaaaa acaataaaaa ctatacaagc ttcccgcctc aagacagaga
     8161 cttga
//
