LOCUS       sequence-209028-        4444 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  knock in bait for Axolotl Tyrosinase locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4444)
  AUTHORS   Fei JF, Schuez M, Tazaki A, Taniguchi Y, Roensch K, Tanaka EM
  TITLE     CRISPR-mediated genomic deletion of Sox2 in the axolotl shows a 
            requirement in spinal cord neural stem cell amplification during 
            tail regeneration.
  JOURNAL   Stem Cell Reports. 2014 Sep 9;3(3):444-59. doi: 
            10.1016/j.stemcr.2014.06.018. Epub 2014 Aug 7.
  PUBMED    25241743
REFERENCE   2  (bases 1 to 4444)
  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: 
            "10.1016/j.stemcr.2014.06.018"; journalName: "Stem Cell Reports"; 
            date: "2014-09-9- 9"; volume: "3"; issue: "3"; pages: "444-59"
COMMENT     Sequence Label: pGEMT-Tyrosinase-bait-T2a-Cherry-pA
FEATURES             Location/Qualifiers
     source          1..4444
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     16..38
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     30..47
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     31..47
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             317..370
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             371..1081
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(516..534)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     784..803
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(1191..1210)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    1256..1311
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(1310..1329)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     promoter        complement(1639..1657)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1640..1657)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1675..1691)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1675..1691)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1688..1710)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1699..1715
                     /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(1723..1729,1730..1747,1748..1753))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1768..1789
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1906..1923)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2077..2665)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2157..2176)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2836..3627,3628..3696))
                     /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     3459..3478
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3697..3801)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3879..4334)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4016..4037)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4228..4247
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgaa ttgtaatacg
       61 actcactata gacgcgtaga acatcgactt cgcgcacgag gcgcccgcct tccttccctg
      121 gcaccggctc ttcctgctgc tctgggagcg ggagatccag aaggtgaccg gcgaccagga
      181 cttcactatc ccctactggg actggaggga cgcccagggc tgcgccgtct gcaccgacca
      241 gctgatgggc gcccggcacc ccaccgtggc caacttactc agcccgggct ccttcttcgc
      301 ttcttggcag gcatgcgagg gcagaggaag tcttctaaca tgcggtgacg tggaggagaa
      361 tcccggccct atggtgagca agggcgagga ggataacatg gccatcatca aggagttcat
      421 gcgcttcaag gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga
      481 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg
      541 ccccctgccc ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta
      601 cgtgaagcac cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa
      661 gtgggagcgc gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc
      721 cctgcaggac ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga
      781 cggccccgta atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc
      841 cgaggacggc gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca
      901 ctacgacgct gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc
      961 ctacaacgtc aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga
     1021 acagtacgaa cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagta
     1081 agaattctgc agtcgagtca tcgcgattaa ttcactcctc aggtgcaggc tgcctatcag
     1141 aaggtggtgg ctggtgtggc caatgccctg gctcacaaat accactgaga tctttttccc
     1201 tctgccaaaa attatgggga catcatgaag ccccttgagc atctgacttc tggctaataa
     1261 aggaaattta ttttcattgc aatagtgtgt tggaattttt tgtgtctctc actcggaagg
     1321 acatatggga gggcaaatca tttaaaacat cagaatgagt atttggttta gagtttggca
     1381 acatatgcca tatgctggct gccatgaaca aaggtggcta taaagaggtc atcagtatat
     1441 gaaacagccc cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat
     1501 tttttttata ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca
     1561 tgttttacta gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct
     1621 tctcttatga agatctattc tatagtgtca cctaaatagc ttggcgtaat catggtcata
     1681 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     1741 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     1801 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     1861 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     1921 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     1981 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     2041 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     2101 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2161 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     2221 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     2281 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     2341 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     2401 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     2461 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     2521 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     2581 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     2641 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     2701 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     2761 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     2821 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     2881 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     2941 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     3001 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     3061 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3121 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3181 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     3241 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     3301 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     3361 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     3421 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     3481 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     3541 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     3601 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     3661 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     3721 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     3781 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgatg cggtgtgaaa
     3841 taccgcacag atgcgtaagg agaaaatacc gcatcaggaa attgtaagcg ttaatatttt
     3901 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     3961 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     4021 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     4081 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     4141 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     4201 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     4261 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     4321 gctacagggc gcgtccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     4381 cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt
     4441 tggg
//
