LOCUS       sequence-346552-        5095 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Leishmania cell free expression of zebrafish Cavin4a-mCherry. Parton
            lab clone GWU.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5095)
  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 5095)
  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-Cavin4a-mCherry
FEATURES             Location/Qualifiers
     source          1..5095
                     /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(1372..1396)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1398..2105
                     /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(1540..1558)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1808..1827
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2002..2025
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    complement(2107..2127)
                     /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(2135..2151)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    2167..2301
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2191..2210)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2245..2264
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2466..2486)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2466..2484)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2505..2521)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2505..2521)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2518..2540)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2529..2545
                     /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(2553..2559,2560..2577,2578..2583))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2598..2619
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2736..2753)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2907..3495)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2987..3006)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3666..4457,4458..4526))
                     /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     4289..4308
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4527..4631)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(join(4657..5095,1..17))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4794..4815)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5006..5025
                     /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 accatggaaa agagaggaga tgtcatatta ggggttgagg
      421 atgaatccgg gcagcccgtc tctgccctgt ccattctgtc tctgctggag cgtgtgtcaa
      481 caatcatcga tggtgtccag gcaagccaac agcgtatgga ggaacgccaa cagcagctcg
      541 agggctcggt cagcgctgtc cagtccgagc ttctcaaact ggctcgtgac cacggggcca
      601 cagcgacgac cgtcgacaaa ctgctgcaga aggcccggcg tgtgagcaca cacgttaaag
      661 aagtacgttc acgtgtggag aaacagaacg tgcgcgtgaa gaaagtggaa actacgcaag
      721 atgagctgct cacacgcaac aagttcaggg ttgtgatcta tcagggtgaa aaagaagtcc
      781 cctctgttgc tgtcaccaag actcccaagg gagcagggct tgctgagttg gaggtggagc
      841 cagatgagta tgacatccct gctgacctgt catctgatga agagtacatg gtggtggagg
      901 atgcggagtc gtccaggggc gcccgcctca aacagtccgg gttaaaaggc atcgagaaca
      961 tcaaggctgc gttctccaaa gagaacatga acaagacccg cgagaagacc cgagaaaacc
     1021 tcagcaagac caaggagagc ctgagtaaga ccggccagac gcttggaacc aaattcaaca
     1081 ctttgggtga gaagatcgtg ccacccgagc agcgagagaa gataaagcag agcagcgaga
     1141 ggctgaagga aaacatcgct aagaaagcgc ccaccaagga atccttcaaa atcaagctga
     1201 agaaggagcg cacggtcgct gaaggccagg agggtgcgga ggcggaacct gctgtgactc
     1261 cacccaaagg caggaagtcc agcccagatg tgacctacac agaggtggtt accgaaaaca
     1321 aaagggaggg tccagtatca gaggagggag caactcgtat ccatgaggac gacccagctt
     1381 tcttgtacaa agtggctgtg agcaagggcg aggaggacaa catggccatc atcaaggagt
     1441 tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg
     1501 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     1561 gcggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg
     1621 cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct
     1681 tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact
     1741 cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct
     1801 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt
     1861 accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg
     1921 gccactacga cgccgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg
     1981 gcgcctacaa cgtcaacatc aagctggaca tcacctccca caacgaggac tacaccatcg
     2041 tggaacagta cgagcgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca
     2101 agtaaccaac tttattatac atagttgata attcactggc cgtcgtttta cggtaccatc
     2161 gatgatgatc cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     2221 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     2281 ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag
     2341 ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg tatggctgat
     2401 tatgatctcg agattaccct gttatcccta gcgtggatcc taggactagt acccagcttt
     2461 tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat catggtcata gctgtttcct
     2521 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccgggag cataaagtgt
     2581 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     2641 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     2701 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     2761 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     2821 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     2881 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     2941 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     3001 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     3061 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     3121 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     3181 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     3241 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     3301 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     3361 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     3421 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     3481 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     3541 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     3601 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     3661 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     3721 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     3781 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     3841 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     3901 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     3961 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     4021 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     4081 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     4141 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     4201 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     4261 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     4321 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     4381 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     4441 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     4501 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     4561 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     4621 ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt
     4681 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     4741 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     4801 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     4861 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     4921 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     4981 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     5041 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgc
//
