LOCUS       sequence-243962-        6928 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent expression of NaChBac (FLAG-tagged) in mammalian 
            cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6928)
  AUTHORS   Fujimoto S, Leiwe MN, Aihara S, Sakaguchi R, Muroyama Y, Kobayakawa 
            R, Kobayakawa K, Saito T, Imai T
  TITLE     Activity-dependent local protection and lateral inhibition control 
            synaptic competition in developing mitral cells in mice.
  JOURNAL   Dev Cell. 2023 Jul 24;58(14):1221-1236.e7. doi: 
            10.1016/j.devcel.2023.05.004. Epub 2023 Jun 7.
  PUBMED    37290446
REFERENCE   2  (bases 1 to 6928)
  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.devcel.2023.05.004"; journalName: "Dev Cell"; date: 
            "2023-07-24- 24"; volume: "58"; issue: "14"; pages: "1221-1236.e7"
COMMENT     Sequence Label: pCALNL-NaChBac
FEATURES             Location/Qualifiers
     source          1..6928
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          139..1155
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1079..1101
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1163..1182
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1230..1263)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1313..2107
                     /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(1367..1386)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1977..1996
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2287..2421
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(2435..2468)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             3336..3359
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3448..3467)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3513..3568
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3567..3586)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3929..3945)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3929..3945)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3942..3964)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3953..3969
                     /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(3977..3983,3984..4001,4002..4007))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4022..4043
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      4128..4262
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4282..4416
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4484..4501)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4655..5243)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4735..4754)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5414..6205,6206..6274))
                     /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     6037..6056
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6275..6379)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        6410..6789
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        join(6791..6928,1..138)
                     /label=chicken beta-actin promoter
ORIGIN
        1 gggcggggcg aggggcgggg cggggcgagg cggagaggtg cggcggcagc caatcagagc
       61 ggcgcgctcc gaaagtttcc ttttatggcg aggcggcggc ggcggcggcc ctataaaaag
      121 cgaagcgcgc ggcgggcggg agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc
      181 gccgcctcgc gccgcccgcc ccggctctga ctgaccgcgt tactcccaca ggtgagcggg
      241 cgggacggcc cttctcctcc gggctgtaat tagcgcttgg tttaatgacg gcttgtttct
      301 tttctgtggc tgcgtgaaag ccttgagggg ctccgggagg gccctttgtg cggggggagc
      361 ggctcggggg gtgcgtgcgt gtgtgtgtgc gtggggagcg ccgcgtgcgg ctccgcgctg
      421 cccggcggct gtgagcgctg cgggcgcggc gcggggcttt gtgcgctccg cagtgtgcgc
      481 gaggggagcg cggccggggg cggtgccccg cggtgcgggg ggggctgcga ggggaacaaa
      541 ggctgcgtgc ggggtgtgtg cgtggggggg tgagcagggg gtgtgggcgc gtcggtcggg
      601 ctgcaacccc ccctgcaccc ccctccccga gttgctgagc acggcccggc ttcgggtgcg
      661 gggctccgta cggggcgtgg cgcggggctc gccgtgccgg gcggggggtg gcggcaggtg
      721 ggggtgccgg gcggggcggg gccgcctcgg gccggggagg gctcggggga ggggcgcggc
      781 ggcccccgga gcgccggcgg ctgtcgaggc gcggcgagcc gcagccattg ccttttatgg
      841 taatcgtgcg agagggcgca gggacttcct ttgtcccaaa tctgtgcgga gccgaaatct
      901 gggaggcgcc gccgcacccc ctctagcggg cgcggggcga agcggtgcgg cgccggcagg
      961 aaggaaatgg gcggggaggg ccttcgtgcg tcgccgcgcc gccgtcccct tctccctctc
     1021 cagcctcggg gctgtccgcg gggggacggc tgccttcggg ggggacgggg cagggcgggg
     1081 ttcggcttct ggcgtgtgac cggcggctct agagcctctg ctaaccatgt tcatgccttc
     1141 ttctttttcc tacagctcct gggcaacgtg ctggttattg tgctgtctca tcattttggc
     1201 aaagaattga ttaattcgag cgaacgcgta taacttcgta tagcatacat tatacgaagt
     1261 tatctcgagt cggatttgat ctgatcaaga gacaggatga ggatcgtttc gcatgattga
     1321 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     1381 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     1441 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga
     1501 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     1561 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     1621 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     1681 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     1741 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     1801 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga
     1861 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     1921 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     1981 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     2041 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     2101 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     2161 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     2221 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     2281 atcgataact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     2341 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     2401 gtatcttatc atgtctggat caatccgaac gcgtataact tcgtatagca tacattatac
     2461 gaagttatct cgagtcgctc ggtacgattt aaattgaatt cgccaccatg aaaatggaag
     2521 ctagacagaa acagaacagt ttcactagta aaatgcaaaa aatcgtgaat catcgggcgt
     2581 ttacgtttac ggtgatcgcg ttaattcttt ttaatgccct tatcgttgga atcgaaacct
     2641 atccacgtat ttatgctgat cataaatggt tattctatcg aatagatctt gttcttctct
     2701 ggatcttcac gattgaaatt gcgatgcgct ttttagcttc gaatccgaaa tcagcttttt
     2761 ttcgaagttc atggaattgg tttgatttct taattgtagc cgcaggtcat atatttgcag
     2821 gtgctcaatt tgtgacggtt ctccgtattt tacgggttct cagggtacta cgggccatct
     2881 cagttgttcc atcgttgcgc aggttagttg atgcgttggt gatgacgatc ccggcgttag
     2941 gaaacatctt aatcttgatg agcattttct tctatatttt tgccgttatc gggacgatgt
     3001 tatttcagca tgtgtcgcct gaatatttcg gtaatttaca gcttagtttg ttaacattgt
     3061 tccaagtggt cacgctagag tcatgggcga gcggcgtcat gcgaccaatt tttgccgaag
     3121 ttccgtggtc ttggctttat tttgtcagct ttgtcttaat cggtacgttt atcatcttta
     3181 acttgtttat cggtgtaatc gtcaataacg ttgaaaaagc agagttaacg gacaatgagg
     3241 aagatggtga agccgatggg ttaaaacaag aaatctcagc cttaagaaaa gacgtagccg
     3301 agctaaaaag cttgcttaaa caattgaaac tcgaggacta caaggacgac gacgacaagt
     3361 gagcggccgc actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa
     3421 tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat
     3481 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat
     3541 agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt
     3601 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata tgctggctgc
     3661 catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc ctgctgtcca
     3721 ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat tttgttttgt
     3781 gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt
     3841 cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga gatccctcga
     3901 cctgcagccc aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     3961 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     4021 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     4081 acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca accatagtcc cgcccctaac
     4141 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     4201 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     4261 cctaggcttt tgcaaaaagc taacttgttt attgcagctt ataatggtta caaataaagc
     4321 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     4381 tccaaactca tcaatgtatc ttatcatgtc tggatccgct gcattaatga atcggccaac
     4441 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     4501 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4561 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4621 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     4681 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4741 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4801 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4861 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4921 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4981 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     5041 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     5101 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     5161 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     5221 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     5281 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     5341 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     5401 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     5461 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     5521 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     5581 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     5641 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     5701 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     5761 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     5821 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     5881 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     5941 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     6001 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     6061 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     6121 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     6181 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     6241 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     6301 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     6361 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgggtcg acattgatta
     6421 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
     6481 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
     6541 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
     6601 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
     6661 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
     6721 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
     6781 attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc ccccccctcc
     6841 ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat gggggcgggg
     6901 gggggggggg ggcgcgccag gcggggcg
//
