LOCUS       sequence-103228-        7526 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector for PhiC31o mRNA synthesis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7526)
  AUTHORS   Ohtsuka M, Miura H, Mochida K, Hirose M, Hasegawa A, Ogura A, 
            Mizutani R, Kimura M, Isotani A, Ikawa M, Sato M, Gurumurthy CB
  TITLE     One-step generation of multiple transgenic mouse lines using an 
            improved Pronuclear Injection-based Targeted Transgenesis (i-PITT).
  JOURNAL   BMC Genomics. 2015 Apr 9;16:274. doi: 10.1186/s12864-015-1432-5.
  PUBMED    25887549
REFERENCE   2  (bases 1 to 7526)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC 
            Genomics."; date: "2015-04-9"; volume: "16:274. doi"; pages: " 
            10.1186/s12864-015-1432-5"
COMMENT     Sequence Label: pBBK
FEATURES             Location/Qualifiers
     source          1..7526
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      949..958
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             955..2769
                     /codon_start=1
                     /product="integrase from phage phi-C31"
                     /label=phage phi-C31 integrase
                     /translation="MDTYAGAYDRQSRERENSSAASPATQRSANEDKAADLQREVERDG
                     GRFRFVGHFSEAPGTSAFGTAERPEFERILNECRAGRLNMIIVYDVSRFSRLKVMDAIP
                     IVSELLALGVTIVSTQEGVFRQGNVMDLIHLIMRLDASHKESSLKSAKILDTKNLQREL
                     GGYVGGKAPYGFELVSETKEITRNGRMVNVVINKLAHSTTPLTGPFEFEPDVIRWWWRE
                     IKTHKHLPFKPGSQAAIHPGSITGLCKRMDADAVPTRGETIGKKTASSAWDPATVMRIL
                     RDPRIAGFAAEVIYKKKPDGTPTTKIEGYRIQRDPITLRPVELDCGPIIEPAEWYELQA
                     WLDGRGRGKGLSRGQAILSAMDKLYCECGAVMTSKRGEESIKDSYRCRRRKVVDPSAPG
                     QHEGTCNVSMAALDKFVAERIFNKIRHAEGDEETLALLWEAARRFGKLTEAPEKSGERA
                     NLVAERADALNALEELYEDRAAGAYDGPVGRKHFRKQQAALTLRQQGAEERLAELEAAE
                     APKLPLDQWFPEDADADPTGPKSWWGRASVDDKRVFVGLFVDKIVVTKSTTGRGQGTPI
                     EKRASITWAKPPTDDDEDDAQDGTEDVAA"
     CDS             2770..2790
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3029..3058
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3074..3091
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3114..3131)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3120..3344
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3390..3818
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3477..3496)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3687..3708
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3827..3847)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3832..4162
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4013..4148
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4075..4094
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4229..5023
                     /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"
     primer_bind     complement(4283..4302)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4893..4912
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5199..5320
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5236..5255)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5290..5309
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5369..5385)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5369..5385)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5382..5404)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5393..5409
                     /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(join(5417..5423,5424..5441,5442..5447))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5462..5483
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5600..5617)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5771..6359)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5851..5870)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6530..7321,7322..7390))
                     /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"
     primer_bind     7153..7172
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7391..7495)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagcttggt accgagctcg gatccactag tccagtgtgg tggaattcgc caccatggat
      961 acctacgccg gagcctacga cagacagagc cgggagagag agaacagcag cgccgccagc
     1021 cccgccaccc agagaagcgc caacgaggat aaggccgccg atctgcagag agaggtggag
     1081 agggacggcg gcagattcag atttgtgggc cacttcagcg aggcccctgg caccagcgcc
     1141 ttcggcaccg ccgagagacc cgagttcgag agaatcctga acgagtgtag ggccggcagg
     1201 ctgaacatga tcatcgtgta cgacgtgtcc cggttcagca ggctgaaggt gatggacgcc
     1261 atccctatcg tgtccgagct gctggccctg ggcgtgacca tcgtgtccac ccaggaaggc
     1321 gtctttagac agggcaacgt gatggacctg atccacctga tcatgaggct ggacgccagc
     1381 cacaaggaga gcagcctgaa gagcgccaag atcctggaca ccaagaacct gcagagggag
     1441 ctgggcggct atgtgggcgg caaggccccc tacggcttcg agctggtgtc cgagaccaag
     1501 gagatcaccc ggaacggcag gatggtgaac gtggtgatca acaagctggc ccacagcacc
     1561 acccccctga ccggcccctt cgagtttgag cccgacgtga tcaggtggtg gtggcgggag
     1621 atcaagaccc acaagcacct gcctttcaag cccggcagcc aggccgccat ccaccccggc
     1681 agcatcaccg gcctgtgtaa gagaatggac gccgacgccg tgcccaccag aggcgagacc
     1741 atcggcaaga aaaccgccag cagcgcctgg gaccccgcca ccgtgatgag aatcctgagg
     1801 gaccctagga tcgccggctt cgccgccgag gtgatctaca agaagaagcc cgacggcacc
     1861 cccaccacca agatcgaggg ctacagaatc cagagagacc ccatcaccct gagacctgtg
     1921 gagctggact gtggccctat catcgagcct gccgagtggt acgagctgca ggcctggctg
     1981 gacggcagag gcagaggcaa gggcctgagc agaggccagg ccatcctgag cgccatggac
     2041 aagctgtact gtgagtgtgg cgccgtgatg accagcaaga gaggcgagga gagcatcaag
     2101 gacagctacc ggtgccggag aagaaaggtg gtggacccca gcgcccctgg ccagcacgag
     2161 ggcacctgta atgtgagcat ggccgccctg gacaagttcg tggccgagcg gatcttcaac
     2221 aagatccggc acgccgaggg cgacgaggag accctggccc tgctgtggga ggccgccaga
     2281 agattcggca agctgaccga ggcccccgag aagagcggcg agagggccaa cctggtggcc
     2341 gagagagccg acgccctgaa cgccctggag gagctgtacg aggacagagc cgccggagcc
     2401 tatgacggcc ctgtgggcag gaagcacttc agaaagcagc aggccgccct gaccctgaga
     2461 cagcagggcg ccgaggaaag actggccgag ctggaggccg ccgaggcccc taagctgccc
     2521 ctggatcagt ggttccccga ggatgccgac gccgacccca ccggccccaa gtcctggtgg
     2581 ggcagagcca gcgtggacga caagagggtg ttcgtgggcc tgttcgtgga taagatcgtg
     2641 gtgaccaaga gcaccaccgg caggggccag ggcaccccca tcgagaagag agccagcatc
     2701 acctgggcca agcctcccac cgacgacgac gaggatgacg cccaggacgg caccgaggac
     2761 gtggccgccc ctaagaaaaa gcggaaagtg tgaattctgc agatatccag cacagtggcg
     2821 gccgctcgac tatgggcacc aaagaacctg taaacgttat ctttttaaat tgaatgtgca
     2881 caaataaaag tttggaaaag aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     2941 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaatacgcat catacgctgt
     3001 ggcgtacctc gagtctagag ggcccttcga acaaaaactc atctcagaag aggatctgaa
     3061 tatgcatacc ggtcatcatc accatcacca ttgagtttaa acccgctgat cagcctcgac
     3121 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     3181 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     3241 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     3301 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     3361 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     3421 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     3481 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     3541 tcggggcatc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     3601 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     3661 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     3721 ccctatctcg gtctattctt ttgatttata agggattttg gggatttcgg cctattggtt
     3781 aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag
     3841 ttagggtgtg gaaagtcccc aggctcccca ggcaggcaga agtatgcaaa gcatgcatct
     3901 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3961 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4021 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4081 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     4141 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     4201 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     4261 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     4321 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     4381 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     4441 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     4501 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     4561 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     4621 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     4681 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     4741 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     4801 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     4861 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     4921 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4981 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     5041 gcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     5101 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     5161 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     5221 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     5281 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     5341 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     5401 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     5461 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     5521 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     5581 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     5641 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5701 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5761 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5821 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5881 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5941 cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt tcggtgtagg
     6001 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6061 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     6121 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     6181 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     6241 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6301 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6361 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6421 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6481 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6541 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6601 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6661 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6721 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6781 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     6841 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     6901 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     6961 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     7021 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     7081 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     7141 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     7201 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     7261 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     7321 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     7381 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7441 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     7501 ttccccgaaa agtgccacct gacgtc
//
