LOCUS       sequence-7055-ex        6868 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6868)
  AUTHORS   Schenten D, Gerlach VL, Guo C, Velasco-Miguel S, Hladik CL, White 
            CL, Friedberg EC, Rajewsky K, Esposito G
  TITLE     DNA polymerase kappa deficiency does not affect somatic 
            hypermutation in mice.
  JOURNAL   Eur J Immunol. 2002 Nov . 32(11):3152-60.
  PUBMED    12555660
REFERENCE   2  (bases 1 to 6868)
  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: "Eur J 
            Immunol. 2002 Nov . 32(11):3152-60."
COMMENT     Sequence Label: pEasy Flox
FEATURES             Location/Qualifiers
     source          1..6868
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    7..40
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(97..120)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     486..505
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             567..1370
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(630..649)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1240..1259
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    2008..2041
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2053..2086
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        2333..2395
                     /label=Mini-TK promoter
                     /note="minimal herpes simplex virus (HSV) thymidine kinase 
                     promoter"
     5'UTR           2417..2456
                     /label=HSV TK 5'-UTR
                     /note="5' untranslated region from the herpes simplex virus
                     thymidine kinase gene"
     CDS             2474..3604
                     /codon_start=1
                     /product="herpes simplex virus thymidine kinase"
                     /label=HSV TK
                     /note="confers sensitivity to nucleoside analogs such as 
                     acyclovir or ganciclovir"
                     /translation="MASYPGHQHASAFDQAARSRGHSNRRTALRPRRQQEATEVRPEQK
                     MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWRVLGASETIANIYTTQ
                     HRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGSSHAPPPALTLIFDR
                     HPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGALPEDRHIDRLAKRQR
                     PGERLDLAMLAAIRRVYGLLANTVRYLQCGGSWREDWGQLSGTAVPPQGAEPQSNAGPR
                     PHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRSMHVFILDYDQSPAGCRDAL
                     LQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
     primer_bind     complement(3602..3621)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3646..3694
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     complement(3984..4003)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3985..4003)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4013..4030)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4013..4029)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4022..4044)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4171..4626
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4258..4277)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4468..4489
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4652..4756
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4757..4825,4826..5617)
                     /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     complement(4975..4994)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5788..6376
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6277..6296
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6530..6547
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6664..6685
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6700..6705,6706..6723,6724..6730)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6738..6754
                     /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)."
     primer_bind     6743..6765
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6762..6778
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6762..6778
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6797..6817
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6799..6817
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 ggatccataa cttcgtataa tgtatgctat acgaagttat ccttaattaa ggaattctac
       61 cgggtagggg aggcgctttt cccaaggcag tctggagcat gcgctttagc agccccgctg
      121 gcacttggcg ctacacaagt ggcctctggc ctcgcacaca ttccacatcc accggtagcg
      181 ccaaccggct ccgttctttg gtggcccctt cgcgccactt ctactcctcc cctagtcagg
      241 aagtttcccc cagcaagctc gcgtcgtgca ggacgtgaca aatggaagta gcacgtctca
      301 ctagtctcgt gcagatggac agcaccgctg agcaatggaa gcgggtaggc ctttggggca
      361 gcggccaata gcagctttgt tccttcgctt tctgggctca gaggctggga aggggtgggt
      421 ccgggggcgg gctcaggggc gggctcaggg gcgggcgggc gcccgaaggt cctcccgagg
      481 cccggcattc tgcacgcttc aaaagcgcac gtctgccgcg ctgttctcct cttcctcatc
      541 tccgggcctt tcgacctgca gccaatatgg gatcggccat tgaacaagat ggattgcacg
      601 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
      661 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
      721 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
      781 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
      841 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
      901 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
      961 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     1021 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     1081 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
     1141 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     1201 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     1261 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     1321 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     1381 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     1441 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     1501 cctccagcgc ggggatctca tgctggagtt cttcgcccac ccccgaaatt gatgatctat
     1561 taaacaataa agatgtccac taaaatggaa gtttttcctg tcatactttg ttaagaaggg
     1621 tgagaacaga gtacctacat tttggaatgg aaggattgga gctacggggg tgggggtggg
     1681 gtgggattag ataaatgcct gctctttact gaaggctctt tactattgct ttatgataat
     1741 gtttcatagt tggatatcat aatttaaaca agcaaaacca aattaagggc cagctcattc
     1801 ctcccactca tgatctatag atctatagat ctctcgtggg atcattgttt ttctcttgat
     1861 tcccacttgt ggttctaagt actgtggttt ccaaatgtgt cagtttcata gcctgaagaa
     1921 cgagatcagc agcctctgtt ccacatacac ttcattctca gtattgtttg gcaagttcta
     1981 attccatcag aagctcctta attaaggata acttcgtata atgtatgcta tacgaagtta
     2041 tctagagtcg acataacttc gtataatgta tgctatacga agttataagc ttgcggccgg
     2101 ccgccacctc gagcagtgtg gttttgcaag aggaagcaaa aagcctctcc acccaggcct
     2161 ggaatgtttc cacccaatgt cgagcagtgt ggttttgcaa gaggaagcaa aaagcctctc
     2221 cacccaggcc tggaatgttt ccacccaatg tcgagcaaac cccgcccagc gtcttgtcat
     2281 tggcgaattc gaacacgcag atgcagtcgg ggcggcgcgg tccgaggtcc acttcgcata
     2341 ttaaggtgac gcgtgtggcc tcgaacaccg agcgaccctg cagcgacccg cttaacagcg
     2401 tcaacagcgt gccgcagatc ttggtggcgt gaaactcccg cacctcttcg gccagcgcct
     2461 tgtagaagcg cgtatggctt cgtaccccgg ccatcaacac gcgtctgcgt tcgaccaggc
     2521 tgcgcgttct cgcggccata gcaaccgacg tacggcgttg cgccctcgcc ggcagcaaga
     2581 agccacggaa gtccgcccgg agcagaaaat gcccacgcta ctgcgggttt atatagacgg
     2641 tccccacggg atggggaaaa ccaccaccac gcaactgctg gtggccctgg gttcgcgcga
     2701 cgatatcgtc tacgtacccg agccgatgac ttactggcgg gtgctggggg cttccgagac
     2761 aatcgcgaac atctacacca cacaacaccg cctcgaccag ggtgagatat cggccgggga
     2821 cgcggcggtg gtaatgacaa gcgcccagat aacaatgggc atgccttatg ccgtgaccga
     2881 cgccgttctg gctcctcata tcggggggga ggctgggagc tcacatgccc cgcccccggc
     2941 cctcaccctc atcttcgacc gccatcccat cgccgccctc ctgtgctacc cggccgcgcg
     3001 gtaccttatg ggcagcatga ccccccaggc cgtgctggcg ttcgtggccc tcatcccgcc
     3061 gaccttgccc ggcaccaaca tcgtgcttgg ggcccttccg gaggacagac acatcgaccg
     3121 cctggccaaa cgccagcgcc ccggcgagcg gctggacctg gctatgctgg ctgcgattcg
     3181 ccgcgtttac gggctacttg ccaatacggt gcggtatctg cagtgcggcg ggtcgtggcg
     3241 ggaggactgg ggacagcttt cggggacggc cgtgccgccc cagggtgccg agccccagag
     3301 caacgcgggc ccacgacccc atatcgggga cacgttattt accctgtttc gggcccccga
     3361 gttgctggcc cccaacggcg acctgtataa cgtgtttgcc tgggccttgg acgtcttggc
     3421 caaacgcctc cgttccatgc acgtctttat cctggattac gaccaatcgc ccgccggctg
     3481 ccgggacgcc ctgctgcaac ttacctccgg gatggtccag acccacgtca ccacccccgg
     3541 ctccataccg acgatatgcg acctggcgcg cacgtttgcc cgggagatgg gggaggctaa
     3601 ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac
     3661 agaataaaac gcacgggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc
     3721 tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc
     3781 ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg
     3841 gcggcaggcc ctgccatagc cactggcccc gtgggttagg gacggggtcc cccatgggga
     3901 atggtttatg gttcgtgggg gttattattt tgggcgttgc gtggggtctg gtggacgacc
     3961 caagctggtg gagctccaat tcgccctata gtgagtcgta ttacgcgcgc tcactggccg
     4021 tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag
     4081 cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     4141 aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     4201 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     4261 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     4321 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     4381 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     4441 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     4501 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     4561 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     4621 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     4681 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     4741 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     4801 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     4861 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     4921 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     4981 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     5041 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     5101 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     5161 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     5221 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     5281 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     5341 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     5401 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     5461 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     5521 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     5581 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     5641 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     5701 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     5761 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     5821 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     5881 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     5941 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     6001 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     6061 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     6121 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     6181 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     6241 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     6301 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     6361 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     6421 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     6481 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     6541 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     6601 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     6661 aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
     6721 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
     6781 tgattacgcc aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg tcgagatcta
     6841 gctagatatc gatggccata gcggccgc
//
