LOCUS       sequence-346527-        5062 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Leishmania cell free expression of zebrafish Cavin4b-mCherry. Parton
            lab clone GWR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5062)
  AUTHORS   Lo HP, Lim YW, Xiong Z, Martel N, Ferguson C, Ariotti N, Giacomotto 
            J, Rae J, Floetenmeyer M, Moradi SV, Gao Y, Tillu VA, Xia D, Wang H,
            Rahnama S, Nixon SJ, Bastiani M, Day RD, Smith KA, Palpant NJ, 
            Johnston WA, Alexandrov K, Collins BM, Hall TE, Parton RG
  TITLE     Cavin4 interacts with Bin1 to promote T-tubule formation and 
            stability in developing skeletal muscle.
  JOURNAL   J Cell Biol. 2021 Dec 6;220(12). pii: 212693. doi: 
            10.1083/jcb.201905065. Epub 2021 Oct 11.
  PUBMED    34633413
REFERENCE   2  (bases 1 to 5062)
  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: "J Cell 
            Biol."; date: "2021-12-6"; volume: "220(12). pii: 212693. doi"; 
            pages: " 10.1083/jcb.201905065. Epub 2021 Oct 11"
COMMENT     Sequence Label: SITS-Cavin4b-mCherry
FEATURES             Location/Qualifiers
     source          1..5062
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    117..137
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     168..187
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        168..186
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             330..353
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     protein_bind    353..377
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      378..387
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(1339..1363)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1365..2072
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1507..1525)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1775..1794
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1969..1992
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    complement(2074..2094)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     complement(2102..2118)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    2134..2268
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2158..2177)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2212..2231
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2433..2453)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2433..2451)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2472..2488)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2472..2488)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2485..2507)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2496..2512
                     /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(2520..2526,2527..2544,2545..2550))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2565..2586
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2703..2720)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2874..3462)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2954..2973)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3633..4424,4425..4493))
                     /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     4256..4275
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4494..4598)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(join(4624..5062,1..17))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4761..4782)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4973..4992
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg aagggcgatc
       61 ggtgcgggcc tcttcgctat tacgccagtc gactagggat aacagggtaa tgaattcaac
      121 tttgtataga aaagttgaca tttccccgaa aagtgccacc tgacgtctaa tacgactcac
      181 tatagggaca tcttaagttt attttatttt attttatttt attttatttt attttatttt
      241 attttatttt attttattta accatgacag taatgtataa agtctgtaaa gacattaaac
      301 acgtaagtga aaccatgggc acctctggtc accaccatca ccaccatcac caccaagttt
      361 gtacaaaaaa gcaggctgcc accatggctg ataaactggg actggcaggt gtggcggacg
      421 agcccagcgc catgaacatc ttctctctgc tggagagggt gtccgggatc atagacaatg
      481 ttcaggcctg ccagcagcgg atggaggaga ggcagctgga gctggagaac accgtcaaga
      541 ccatccaagc agatgtagtc aagctgacga aggagcacac ggcgaccagt ggcaccgtgg
      601 agaaacttct ggagaaaaca cgcaaggtca gctgccacat caaggacgtc cgcgtgcggg
      661 tggagaagca gaacgttcga gtcaagaagg tggaggaaac ccaagtcgag ctgctgaata
      721 agaacaagtt ccgtgtcgtc atctaccagg gagataatga ggtcccagct gtggctggtc
      781 ctaaagcctc tccgaaggga gcaatgggtg gagacgctgc tgcagaccca gatgcaccag
      841 aggctccgct ccctgactcg gatgaggaat acatggttgt tgaagaagcc gattcttcca
      901 ctgctgccaa actaaagaag accggcctga agcgcatcga aagcctgaag cagaccttct
      961 cacgcgagaa catgactaag accaaagaaa acctgggcac caaggtcaac aagctgggcg
     1021 agcgcatagt aacagccgaa cggcgtgaga aaatccgcca gtcaggagaa cgactgaaac
     1081 agtctggcga gcgcttcaag gaaaccatca caaaaaccgt accagcaaag ctgaatctga
     1141 agaaggaaag gacggtggct gagggtcagg agggggcaga agggaccaca gaaggaacag
     1201 cacccgtccc accaccgaag ggtcgtaaac ccagtcctga ggtggcctac acagagccaa
     1261 cgcagaaaca agaaggtcca gatgagggca agggtgagga atcagaagta ccgatgtatg
     1321 atatgaagca gctttctgac ccagctttct tgtacaaagt ggctgtgagc aagggcgagg
     1381 aggacaacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg
     1441 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc
     1501 agaccgccaa gctgaaggtg accaagggcg gccccctgcc cttcgcctgg gacatcctgt
     1561 cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac atccccgact
     1621 acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg
     1681 gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggcgagttc atctacaagg
     1741 tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag aagaccatgg
     1801 gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag ggcgagatca
     1861 agcagaggct gaagctgaag gacggcggcc actacgacgc cgaggtcaag accacctaca
     1921 aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag ctggacatca
     1981 cctcccacaa cgaggactac accatcgtgg aacagtacga gcgcgccgag ggccgccact
     2041 ccaccggcgg catggacgag ctgtacaagt aaccaacttt attatacata gttgataatt
     2101 cactggccgt cgttttacgg taccatcgat gatgatccag acatgataag atacattgat
     2161 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
     2221 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     2281 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa
     2341 aacctctaca aatgtggtat ggctgattat gatctcgaga ttaccctgtt atccctagcg
     2401 tggatcctag gactagtacc cagcttttgt tccctttagt gagggttaat tgcgcgcttg
     2461 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     2521 aacatacgag ccgggagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     2581 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     2641 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     2701 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     2761 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     2821 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     2881 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     2941 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3001 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3061 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3121 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3181 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3241 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     3301 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     3361 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     3421 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     3481 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     3541 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     3601 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     3661 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     3721 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     3781 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     3841 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     3901 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     3961 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4021 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     4081 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     4141 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     4201 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     4261 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     4321 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     4381 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     4441 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     4501 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     4561 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     4621 cctaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     4681 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     4741 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     4801 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     4861 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     4921 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     4981 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     5041 ccacacccgc cgcgcttaat gc
//
