LOCUS       sequence-79584-e        3459 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To generate transgenic axolotls using Tol2 or SceI mediated 
            transgenics.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3459)
  AUTHORS   Khattak S, Murawala P, Andreas H, Kappert V, Schuez M, 
            Sandoval-Guzman T, Crawford K, Tanaka EM
  TITLE     Optimized axolotl (Ambystoma mexicanum) husbandry, breeding, 
            metamorphosis, transgenesis and tamoxifen-mediated recombination.
  JOURNAL   Nat Protoc. 2014 Mar;9(3):529-40. doi: 10.1038/nprot.2014.040. Epub 
            2014 Feb 6.
  PUBMED    24504478
REFERENCE   2  (bases 1 to 3459)
  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"; doi: 
            "10.1038/nprot.2014.040"; journalName: "Nat Protoc"; date: 
            "2014-03"; volume: "9"; issue: "3"; pages: "529-40"
COMMENT     Sequence Label: pBSII-SK-mTol2-MCS
FEATURES             Location/Qualifiers
     source          1..3459
                     /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"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(659..670,671..858)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     promoter        877..895
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     947..963
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     947..963
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(997..1013)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(998..1014)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(1056..1074)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(1095..1232,1233..1244)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     promoter        complement(1271..1289)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1310..1326)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1310..1326)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1323..1345)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1334..1350
                     /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(1358..1364,1365..1382,1383..1388))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1403..1424
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1541..1558)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1712..2300)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1792..1811)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2471..3262,3263..3331))
                     /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     3094..3113
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3332..3436)
                     /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 gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtaccca
      661 gaggtgtaaa gtacttgagt aattttactt gattactgta cttaagtatt atttttgggg
      721 atttttactt tacttgagta caattaaaaa tcaatacttt tacttttact taattacatt
      781 tttttagaaa aaaaagtact ttttactcct tacaatttta tttacagtca aaaagtactt
      841 attttttgga gatcacttgg gcccggctcg acgcgcaatt aaccctcact aaagggaaca
      901 aaagctggag cttagggata acagggtaat ccgcggtggc ggccgctcta gaactagtgg
      961 atcccccggg ctgcaggaat tcgatatcaa gcttatcgat accgtcgacc tcgagggggg
     1021 gcccattacc ctgttatccc tagtacccaa ttcgccctat agtgagtcgt attacgcgcg
     1081 tcgagattag atctaatact caagtacaat tttaatggag tactttttta cttttactca
     1141 agtaagattc tagccagata cttttacttt taattgagta aaattttccc taagtacttg
     1201 tactttcact tgagtaaaat ttttgagtac tttttacacc tctgccgcgg tggagctcca
     1261 gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg tcatagctgt
     1321 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     1381 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     1441 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     1501 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     1561 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     1621 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     1681 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     1741 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     1801 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     1861 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     1921 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     1981 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     2041 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     2101 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     2161 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     2221 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     2281 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     2341 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     2401 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     2461 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     2521 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     2581 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     2641 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     2701 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     2761 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     2821 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     2881 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     2941 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     3001 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     3061 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     3121 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     3181 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     3241 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     3301 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     3361 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     3421 aaataggggt tccgcgcaca tttccccgaa aagtgccac
//
