LOCUS       pNWA269_(pCMV6-A        7181 bp DNA     circular SYN 05-SEP-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7181)
  AUTHORS   nichd_dir_cssc_sub
  TITLE     Direct Submission
  JOURNAL   Exported Apr 24, 2023 from SnapGene 6.2.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7181
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     166..182
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        343..722
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        723..926
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        952..970
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1014..1023
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1023..1046
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1065..2153
                     /codon_start=1
                     /label=1-363 RAD23B
                     /translation="MQVTLKTLQQQTFKIDIDPEETVKALKEKIESEKGKDAFPVAGQK
                     LIYAGKILNDDTALKEYKIDEKNFVVVMVTKPKAVSTPAPATTQQSAPASTTAVTSSTT
                     TTVAQAPTPVPALAPTSTPASITPASATASSEPAPASAAKQEKPAEKPAETPVATSPTA
                     TDSTSGDSSRSNLFEDATSALVTGQSYENMVTEIMSMGYEREQVIAALRASFNNPDRAV
                     EYLLMGIPGDRESQAVVDPPQAASTGVPQSSAVAAAAATTTATTTTTSSGGHPLEFLRN
                     QPQFQQMRQIIQQNPSLLPALLQQIGRENPQLLQQISQHQEHFIQMLNEPVQEAGGQGG
                     GGGGGSGGIAEAGSGHMNYIQVT"
     misc_feature    1788..2196
                     /label=Synthetic fragment
                     /note="Cloned into Bsu36I-MfeI of pNWA41"
     misc_feature    2175..2312
                     /label=364-409 RAD23B CDS
     primer_bind     complement(2389..2405)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    2416..3038
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     promoter        3067..3396
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3247..3382
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3463..4257
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    4431..4552
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4601..4617)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4625..4641
                     /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(4649..4679)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4694..4715
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5003..5588)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5759..6619)
                     /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"
     promoter        complement(6620..6724)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6751..7179
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 aacaaaatat taacgcttac aatttccatt cgccattcag gctgcgcaac tgttgggaag
       61 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
      121 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
      181 gtgccaagct gatctataca ttgaatcaat attggcaatt agccatatta gtcattggtt
      241 atatagcata aatcaatatt ggctattggc cattgcatac gttgtatcta tatcataata
      301 tgtacattta tattggctca tgtccaatat gaccgccatg ttgacattga ttattgacta
      361 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
      421 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      481 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
      541 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
      601 gtccgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
      661 tgaccttacg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
      721 tggtgatgcg gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat
      781 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
      841 actttccaaa atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac
      901 ggtgggaggt ctatataagc agagctcgtt tagtgaaccg tcagaatttt gtaatacgac
      961 tcactatagg gcggccggga attcgtcgac tggatccggt accgaggaga tctgccacca
     1021 tggattacaa ggatgacgac gataaggcga tcgccatgga gcatatgcag gtgacactga
     1081 agacactgca gcagcagact tttaagattg acattgaccc cgaggagacc gtgaaggcac
     1141 tgaaggagaa aatcgagagc gaaaagggca aagacgcatt ccctgtggcc gggcagaagc
     1201 tgatctacgc cggaaaaatt ctgaacgacg ataccgctct gaaggaatac aagatcgatg
     1261 agaagaattt cgtggtcgtg atggtcacta agcctaaagc cgtgtccacc ccagctcccg
     1321 caaccacaca gcagtctgcc ccagctagta ctaccgccgt gacaagctcc acaactacca
     1381 cagtcgcaca ggcacctact ccagtgccag ctctggcacc cacatcaact cctgccagca
     1441 tcacaccagc cagcgctact gcatctagtg aacctgcccc agcttccgcc gctaagcagg
     1501 agaaaccagc tgaaaaaccc gcagagactc ctgtggccac ctcacccacc gctacagaca
     1561 gcacctccgg cgattcaagc cggtccaacc tgttcgagga cgctacctcc gcactggtca
     1621 caggccagtc ttacgaaaat atggtgacag agatcatgtc tatggggtat gagcgggaac
     1681 aggtcattgc agccctgaga gctagtttta acaatcccga tagggcagtg gaatacctgc
     1741 tgatgggcat ccctggggac agagagtctc aggcagtcgt ggacccccct caggctgcaa
     1801 gtaccggagt cccacagtcc tctgcagtgg cagctgcagc agctactacc acagctacta
     1861 ccacaactac cagttcaggc gggcaccctc tggagttcct gcgcaaccag ccacagtttc
     1921 agcagatgcg acagatcatt cagcagaatc catccctgct gccagcactg ctgcagcaga
     1981 ttggacgcga aaacccccag ctgctgcagc agatctctca gcaccaggag catttcattc
     2041 agatgctgaa tgaacctgtg caggaggcag gaggacaggg aggaggagga ggaggcggga
     2101 gtggaggcat cgcagaggcc gggtcaggac acatgaacta catccaggtc acataagcga
     2161 tcgccatgga gcatccacag gagaaggaag caattgaaag gctgaaagcc ctgggattcc
     2221 ccgagggcct ggtcatccag gcctattttg cttgcgagaa gaacgaaaat ctggcagcca
     2281 acttcctgct gcagcagaat tttgacgagg attaagaatt cggatccacg cgtaagcttc
     2341 tcgagacgcg ttaagcggcc gcactcgagg tttaaacggc cggccgcggt catagctgtt
     2401 tcctgaacag atcccgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg
     2461 gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg
     2521 tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg
     2581 caagttggga agacaacctg tagggcctgc ggggtctatt gggaaccaag ctggagtgca
     2641 gtggcacaat cttggctcac tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct
     2701 cagcctcccg agttgttggg attccaggca tgcatgacca ggctcagcta atttttgttt
     2761 ttttggtaga gacggggttt caccatattg gccaggctgg tctccaactc ctaatctcag
     2821 gtgatctacc caccttggcc tcccaaattg ctgggattac aggcgtgaac cactgctccc
     2881 ttccctgtcc ttctgatttt aaaataacta taccagcagg aggacgtcca gacacagcat
     2941 aggctacctg gccatgccca accggtggga catttgagtt gcttgcttgg cactgtcctc
     3001 tcatgcgttg ggtccactca gtagatgcct gttgaattgg gtacgcggcc agcttggctg
     3061 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3121 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3181 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3241 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3301 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     3361 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     3421 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     3481 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     3541 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3601 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     3661 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     3721 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     3781 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     3841 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     3901 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     3961 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     4021 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     4081 atcgactgtg gccggctggg tgtggccgac cgctatcagg acatagcgtt ggctacccgt
     4141 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4201 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4261 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4321 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4381 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     4441 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     4501 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     4561 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     4621 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     4681 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     4741 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     4801 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4861 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4921 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4981 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     5041 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     5101 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     5161 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     5221 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     5281 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     5341 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     5401 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     5461 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     5521 aaaccaccgc tggtagcggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5581 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5641 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     5701 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     5761 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     5821 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     5881 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     5941 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6001 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6061 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6121 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6181 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6241 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6301 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6361 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6421 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6481 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     6541 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     6601 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     6661 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     6721 cgcgcacatt tccccgaaaa gtgccacctg acgcgccctg tagcggcgca ttaagcgcgg
     6781 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     6841 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     6901 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     6961 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     7021 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     7081 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     7141 taaaaaatga gctgatttaa caaaaattta acgcgaattt t
//
