LOCUS       sequence-372878-        7441 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses mCherry-Climp63 with partial luminal domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7441)
  AUTHORS   Wang B, Zhao Z, Xiong M, Yan R, Xu K
  TITLE     The endoplasmic reticulum adopts two distinct tubule forms.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 May 3;119(18):e2117559119. doi: 
            10.1073/pnas.2117559119. Epub 2022 Apr 26.
  PUBMED    35471903
REFERENCE   2  (bases 1 to 7441)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl 
            Acad Sci U S A."; date: "2022-05-3"; volume: "119(18):e2117559119. 
            doi"; pages: " 10.1073/pnas.2117559119. Epub 2022 Apr 26"
COMMENT     Sequence Label: mCherry-Climp63(1–437)
FEATURES             Location/Qualifiers
     source          1..7441
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             958..1665
                     /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(1103..1121)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1371..1390
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3038..3262
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3308..3736
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3395..3414)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3605..3626
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3745..3765)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3750..4079
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3930..4065
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3992..4011
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4146..4940
                     /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(4200..4219)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4810..4829
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5114..5235
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5151..5170)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5205..5224
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5284..5300)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5284..5300)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5297..5319)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5308..5324
                     /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(5332..5338,5339..5356,5357..5362))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5377..5398
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5515..5532)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5686..6274)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5766..5785)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6445..7236,7237..7305))
                     /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     7068..7087
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7306..7410)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattcatg
      961 gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg
     1021 cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc
     1081 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc
     1141 gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc
     1201 gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg
     1261 atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc
     1321 gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg
     1381 cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc
     1441 ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag
     1501 gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac
     1561 atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc
     1621 gccgagggcc gccactccac cggcggcatg gacgagctgt acaagtccgg actcagatct
     1681 cgagctccct cggccaaaca aaggggctcc aagggcggcc acggcgccgc gagcccctcg
     1741 gacaagggcg cccacccgtc gggcggcgcg gatgacgtgg cgaagaagcc gccggcggcg
     1801 ccgcagcagc cgcagcctcc cgcgccgcac ccgccgcagc acccgcagaa ccaggcgcac
     1861 cggggcggcc accgcggccg gtcctcagcc gccaccgcca acgcgtcctc cgcgtcctgc
     1921 tcgcgcaggc ttggccgggt gctcaacttt ctcttctacc tctccctggt ggcggcggcc
     1981 gccttctcgg gctggtatgt ccatcacgtc ctggaggagg tccagcaggt ccggcgtggc
     2041 caccaggact tctcccgtca gagggatgag ctgggccagg gcttgcaggg cgtcgagcag
     2101 aaggtccaat ccctgcaagc cacatttggg actttcgagt ccctcctgag aaactcccag
     2161 cataaacagg atctcacaga gaaagccgtg aaggaagggg agagcgagct caaccgaatt
     2221 agcgaagttc tacagaagtt gcagaatgag attctcaaag atctgtcgga cgggatccac
     2281 gtggtgaagg atgcccggga gcgggacttc acgtccctgg agaacacagt cgaggagcgc
     2341 ctgacggagc ttaccaagtc tatcaatgac aacatcgcca tcttcactga tgtccagaag
     2401 aggagccaga aggagatcaa tgaggtcaag atgaaggtgg cctccctgga agaatccaag
     2461 ggggatcgca gtcaggacgt gaaaactctg aaggacgccg tgaaggaggt ccaggcctcc
     2521 atgatgtcga gggagaggga cattgaggcc ctgaagagtt cccttcagac gatggagtcc
     2581 gacgtctaca cagaggtgcg ggaactggtg agcctcaagc aggagcaaca ggcgttcaag
     2641 caggcggccg actcagagcg cctggctctg caggccctca cggagaagct tctgcggtcc
     2701 gaggagtcct cctctcgtct cccagaggac atccggagac tggaggaaga gctacagcag
     2761 ctgaaggtcg gggcccatgg gtcagaggaa ggtgccgtct tcaaagactc caaagcctta
     2821 gaggaactgc agaggcagat tgagggcctg ggtgccaggc tccagtatgt agaggacggg
     2881 gtctactcaa tgcaggtggc ttctgcacgc cacaccgaaa gcctggagtc gctcctgtcc
     2941 aagagccagg agtacgagca acgcctggct atgctgcagg agcacgtggg aaacctgtaa
     3001 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
     3061 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     3121 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     3181 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     3241 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     3301 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     3361 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     3421 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     3481 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     3541 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     3601 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     3661 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     3721 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     3781 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     3841 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3901 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     3961 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     4021 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     4081 agctcccggg agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt
     4141 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     4201 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     4261 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     4321 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     4381 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     4441 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     4501 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     4561 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     4621 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     4681 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     4741 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     4801 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     4861 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     4921 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     4981 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     5041 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     5101 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     5161 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     5221 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     5281 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     5341 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     5401 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     5461 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     5521 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     5581 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     5641 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     5701 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5761 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5821 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     5881 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     5941 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     6001 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     6061 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     6121 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     6181 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     6241 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     6301 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     6361 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     6421 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     6481 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     6541 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     6601 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     6661 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     6721 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     6781 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     6841 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     6901 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     6961 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     7021 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     7081 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     7141 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     7201 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     7261 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     7321 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     7381 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     7441 c
//
