LOCUS       sequence-337468-        4796 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  lox2272 (lox variant)-flanked pPgk-pEM7-Neo*-bpA positive selection 
            cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4796)
  AUTHORS   Nam HS, Capecchi MR
  TITLE     Additional gene targeting vector construction plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4796)
  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: 
            "Unpublished"
COMMENT     Sequence Label: lox2272 Neo* lox2272
FEATURES             Location/Qualifiers
     source          1..4796
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     623..642
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        623..641
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     666..682
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     667..683
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    705..738
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     promoter        802..1301
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(837..860)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1234..1253
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        1313..1360
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             1379..2182
                     /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
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1442..1461)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2052..2071
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2214..2231)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2220..2444
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    2502..2535
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     primer_bind     complement(2555..2571)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2555..2571)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(2608..2626)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2647..2663)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2647..2663)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2660..2682)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2671..2687
                     /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(2695..2701,2702..2719,2720..2725))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2740..2761
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2878..2895)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3049..3637)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3129..3148)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3808..4599,4600..4668))
                     /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     4431..4450
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4669..4773)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgaat tgtaatacga ctcactatag ggcgaattgg gtaccgggcc
      661 ccccctcgag gtcgacggta tcgataagct tgatatcgaa ttccataact tcgtataaag
      721 tatcctatac gaagttatag gtctgaagag gagtttacgt ccagccaagc tagcttggct
      781 gcaggtcgtc gaaattctac cgggtagggg aggcgctttt cccaaggcag tctggagcat
      841 gcgctttagc agccccgctg ggcacttggc gctacacaag tggcctctgg cctcgcacac
      901 attccacatc caccggtagg cgccaaccgg ctccgttctt tggtggcccc ttcgcgccac
      961 cttctactcc tcccctagtc aggaagttcc cccccgcccc gcagctcgcg tcgtgcagga
     1021 cgtgacaaat ggaagtagca cgtctcacta gtctcgtgca gatggacagc accgctgagc
     1081 aatggaagcg ggtaggcctt tggggcagcg gccaatagca gctttgctcc ttcgctttct
     1141 gggctcagag gctgggaagg ggtgggtccg ggggcgggct caggggcggg ctcaggggcg
     1201 gggcgggcgc ccgaaggtcc tccggaggcc cggcattctg cacgcttcaa aagcgcacgt
     1261 ctgccgcgct gttctcctct tcctcatctc cgggcctttc gacctgcagc ctgttgacaa
     1321 ttaatcatcg gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat
     1381 gggatcggcc attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     1441 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     1501 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     1561 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     1621 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     1681 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     1741 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     1801 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     1861 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     1921 gcccgacggc gatgatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     1981 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     2041 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     2101 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     2161 ccttcttgac gagttcttct gaggggatca attctctaga gctcgctgat cagcctcgac
     2221 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     2281 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     2341 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     2401 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     2461 aaccagctgg ggctcgacta gagcttgcgg aacccttcga aataacttcg tataaagtat
     2521 cctatacgaa gttatatcag tcaggtacat aatggatcca ctagttctag agcggccgcc
     2581 accgcggtgg agctccagct tttgttccct ttagtgaggg ttaatttcga gcttggcgta
     2641 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     2701 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     2761 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     2821 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     2881 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     2941 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3001 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3061 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3121 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3181 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3241 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3301 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3361 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3421 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3481 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3541 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3601 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3661 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     3721 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     3781 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     3841 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     3901 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     3961 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     4021 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     4081 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     4141 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     4201 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     4261 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     4321 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     4381 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     4441 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     4501 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     4561 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     4621 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     4681 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     4741 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccac
//
