LOCUS       sequence-451929-        5296 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Encodes mutant siRNA resistant SigmaR1 protein (substitution) 
            labeled with mNeonGreen.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5296)
  AUTHORS   Sawyer EM, Jensen LE, Meehl JB, Larsen KP, Petito DA, Hurley JH, 
            Voeltz GK
  TITLE     SigmaR1 shapes rough endoplasmic reticulum membrane sheets.
  JOURNAL   Dev Cell. 2024 Jul 2:S1534-5807(24)00382-4. doi: 
            10.1016/j.devcel.2024.06.005.
  PUBMED    38971154
REFERENCE   2  (bases 1 to 5296)
  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: "Dev Cell. 
            2024 Jul 2:S1534-5807(24)00382-4. doi: 
            10.1016/j.devcel.2024.06.005."
COMMENT     Sequence Label: siRNA-resistant SigmaR1-mNeonGreen R119A
FEATURES             Location/Qualifiers
     source          1..5296
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      202..211
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      463..472
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(1222..1425)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     enhancer        1426..1729
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(1904..2492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1984..2003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    2821..2868
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     2893..2912
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3100..3894)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(3100..3894)
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3211..3230)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3821..3840
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(3929..4125)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        complement(3937..4147)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3943..4078
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4171..4191
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(4216..4320)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4347..4802
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4434..4453)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4644..4665
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4809..4930
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4820..4839)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4874..4893
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(5052..5296,1..466))
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
ORIGIN
        1 ggtaagtctt cttcgacctg caccagtccg cagcggtcag cgagttggtc atcacaggac
       61 cgtcagcagg gaaaccagtc cccttcacct gggcctctcc tttgatgtgg cttccctcgt
      121 aggtgtagcg gtagttaaca gtaagggagg caccatcttc aaactgcatt gtgcgatgga
      181 cttggtagcc ggagccatct accatggcgg cctggaaagg cgacatcccg tcagggtagg
      241 gcaggtactg atggaagcca tacccgatat gagggaccag aatccagggg gagaactgga
      301 ggtcaccctt ggtggacttc aggtttaact cctcataacc atcatttgga ttgccggtgc
      361 cctgacccac catgtcaaag tccacaccgt tgatggagcc aaagatgtgt aactcatgtg
      421 tcgctgggag agaggccatg ttatcctcct cgcccttgct caccatggtg gcgaccggtg
      481 gatcccgggc ccgcggtacc ctagggtcct ggccaaagag gtatgtcgtc aattccaagc
      541 ggaggccccg agcataggag cgaagagtat agaagagggt gaggaagtcc tgggtgctga
      601 agacagtgtc ggccagcgcg aaggccaggg tggatgggat gacgccccgg ccgtactcga
      661 ccatccaagt attgggcccc cactccacag ctgttgcctc accaggcccg tgtactaccg
      721 tctcccctgg gtagaagacc tcacttttgg tggtgccctc tctccactgg tggaaggtgc
      781 cagagatgat ggtatccgag atctcagccc agtaagcccc cgagtggccg cgggagccca
      841 aggcggtgcc gaagagcagc acatactcgg acagcgaggc gtgcagaagg cacatggcgc
      901 ccatccatcc accagcgttg acgaacaccc actgcagctc ctcgtcgggc agcacgtggc
      961 ctgggtgcag ccgccgcagc tccacgatca gtctgctaaa tgcgagttcg tggtccagcc
     1021 cagcgtactg ccgcgccaac tgcgctatct cttcgcgctg gaagacgaag ctctgcgtac
     1081 ccagccagag ccagacgacc tgggtcagca ccgctgcgac agccaggagc agcgcggccc
     1141 acgcccaccg ccggcccacg gcccactgca tctcgagatc tgagtccggt agcgctagcg
     1201 gatctgacgg ttcactaaac cagctctgct tatatagacc tcccaccgta cacgcctacc
     1261 gcccatttgc gtcaatgggg cggagttgtt acgacatttt ggaaagtccc gttgattttg
     1321 gtgccaaaac aaactcccat tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc
     1381 aaaccgctat ccacgcccat tgatgtactg ccaaaaccgc atcaccatgg taatagcgat
     1441 gactaatacg tagatgtact gccaagtagg aaagtcccat aaggtcatgt actgggcata
     1501 atgccaggcg ggccatttac cgtcattgac gtcaataggg ggcgtacttg gcatatgata
     1561 cacttgatgt actgccaagt gggcagttta ccgtaaatac tccacccatt gacgtcaatg
     1621 gaaagtccct attggcgtta ctatgggaac atacgtcatt attgacgtca atgggcgggg
     1681 gtcgttgggc ggtcagccag gcgggccatt taccgtaagt tatgtaacgc ggaactccat
     1741 atatgggcta tgaactaatg accccgtaat tgattactat taataactaa tgcatggcgg
     1801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     1921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     1981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2461 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2641 atgagtaacc tgaggctatg gcagggcctg ccgccccgac gttggctgcg agccctgggc
     2701 cttcacccga acttgggggg tggggtgggg aaaaggaaga aacgcgggcg tattggcccc
     2761 aatggggtct cggtggggta tcgacagagt gccagccctg ggaccgaacc ccgcgtttat
     2821 gaacaaacga cccaacaccg tgcgttttat tctgtctttt tattgccgtc atagcgcggg
     2881 ttccttccgg tattgtctcc ttccgtgttt cagttagcct ccccctaggg tgggcgaaga
     2941 actccagcat gagatccccg cgctggagga tcatccagcc ggcgtcccgg aaaacgattc
     3001 cgaagcccaa cctttcatag aaggcggcgg tggaatcgaa atctcgtgat ggcaggttgg
     3061 gcgtcgcttg gtcggtcatt tcgaacccca gagtcccgct cagaagaact cgtcaagaag
     3121 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg
     3181 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg
     3241 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc
     3301 caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct cgccgtcggg
     3361 catgctcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc
     3421 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg
     3481 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc
     3541 atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc
     3601 cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc
     3661 tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cttgcagttc
     3721 attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag
     3781 ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag
     3841 cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatgcgaaa
     3901 cgatcctcat cctgtctctt gatcgatctt tgcaaaagcc taggcctcca aaaaagcctc
     3961 ctcactactt ctggaatagc tcagaggccg aggcggcctc ggcctctgca taaataaaaa
     4021 aaattagtca gccatggggc ggagaatggg cggaactggg cggagttagg ggcgggatgg
     4081 gcggagttag gggcgggact atggttgctg actaattgag atgcatgctt tgcatacttc
     4141 tgcctgctgg ggagcctggg gactttccac accctaactg acacacattc cacagctggt
     4201 tctttccgcc tcaggactct tcctttttca atattattga agcatttatc agggttattg
     4261 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     4321 cacatttccc cgaaaagtgc cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg
     4381 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     4441 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     4501 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     4561 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     4621 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     4681 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     4741 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
     4801 ttacgcctta agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     4861 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     4921 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     4981 ggaggttttt taaagcaagt aaaacctcta caaatgtggt atggctgatt atgatctaga
     5041 gtcgcggccg cttacttgta cagctcgtcc atgcccatca catcggtaaa ggccttttgc
     5101 cactccttga agttgagctc ggtcttggag tgcttgagct ccgtcttacg gaacacgtac
     5161 atcggctggt tcttcagata gttagccgcc attggcttgg caaaggtgta ggtggtccgc
     5221 gcagtgctcc ggtagcgctt gccatttcca gtggtgtaac tccacttaaa ggtactgatg
     5281 atggttttgt cgttgg
//
