LOCUS       sequence-434106-        5532 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Truncated Nir1-LNS2 domain lacking N-terminal amphipathic helix; 
            loses ability to bind PA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5532)
  AUTHORS   Weckerly CC, Rahn TA, Ehrlich M, Wills RC, Pemberton JG, Airola MV, 
            Hammond GRV
  TITLE     PILS-Nir1 is a sensitive phosphatidic acid biosensor that reveals 
            mechanisms of lipid production.
  JOURNAL   J Cell Biol. 2025 Nov 3;224(11):e202405174. doi: 
            10.1083/jcb.202405174. Epub 2025 Sep 9.
  PUBMED    40923975
REFERENCE   2  (bases 1 to 5532)
  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: "2025-11-3"; volume: "224(11):e202405174. doi"; pages:
            " 10.1083/jcb.202405174. Epub 2025 Sep 9"
COMMENT     Sequence Label: pNeonGreen-C1-Nir1-631-894
FEATURES             Location/Qualifiers
     source          1..5532
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1320
                     /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"
     regulatory      868..877
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    2131..2196
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2320..2441
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2357..2376)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2411..2430
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2448..2903)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2585..2606)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2797..2816
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2930..3034
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3036..3393
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3036..3227
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3059..3079)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3244..3379
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3428..4222
                     /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             3428..4222
                     /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(3482..3501)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4092..4111
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4410..4429)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4454..4501
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4830..5418
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5319..5338
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gaggataaca tggcctctct cccagcgaca
      661 catgagttac acatctttgg ctccatcaac ggtgtggact ttgacatggt gggtcagggc
      721 accggcaatc caaatgatgg ttatgaggag ttaaacctga agtccaccaa gggtgacctc
      781 cagttctccc cctggattct ggtccctcat atcgggtatg gcttccatca gtacctgccc
      841 taccctgacg ggatgtcgcc tttccaggcc gccatggtag atggctccgg ataccaagtc
      901 catcgcacaa tgcagtttga agatggtgcc tcccttactg ttaactaccg ctacacctac
      961 gagggaagcc acatcaaagg agaggcccag gtgaagggga ctggtttccc tgctgacggt
     1021 cctgtgatga ccaactcgct gaccgctgcg gactggtgca ggtcgaagaa gacttacccc
     1081 aacgacaaaa ccatcatcag tacctttaag tggagttaca ccactggaaa tggcaagcgc
     1141 taccggagca ctgcgcggac cacctacacc tttgccaagc caatggcggc taactatctg
     1201 aagaaccagc cgatgtacgt gttccgtaag acggagctca agcactccaa gaccgagctc
     1261 aacttcaagg agtggcaaaa ggcctttacc gatgtgatgg gcatggacga gctgtacaag
     1321 ggaggctcag gaggcaacgt taccgctaac caccgtgcga acgatgttat cgcggcggaa
     1381 gatggtccgc aggttctggt tggtcgtttc atgtacggtc cgctggacat ggttgctctg
     1441 accggtgaaa aagtggatat cctggttatg gcagaaccgt ctagcggtcg ctgggttcac
     1501 ctggacactg aaatcaccaa ctcctctggt cgtatcacct acaacgttcc gcgtccgcgc
     1561 cgcctgggtg ttggtgttta cccggttaaa atggttgttc gtggtgacca gacctgcgcg
     1621 atgtcttacc tgaccgttct gccgcgcggt atggaatgtg ttgtgttctc tatcgacggt
     1681 tctttcgctg cgagcgtttc tatcatgggc tctgacccga aagttcgtcc gggtgcggtt
     1741 gacgttgttc gtcactggca ggacctgggt tacatgatcc tgtacatcac cggccgtccg
     1801 gacatgcaga aacagcgtgt tgtttcttgg ctgtctcagc acaacttccc gcagggtatg
     1861 atcttcttct ctgatggtct ggttcacgac ccgctgcgtc agaaagcgat cttcctgcgt
     1921 aacctgatgc aggaatgctt catcaaaatc tctgcggcgt acggttctac caaagacatc
     1981 tctgtttact ccgttctggg tctgccggct tctcagatct tcatcgtggg tcgtccgacc
     2041 aaaaaatacc agacccagtg tcagttcctg tctgaaggtt acgcggcgca cctggctgtt
     2101 ctggaagctt ctcaccgttc tcgtccgtaa tccggactca gatctcgagc tcaagcttcg
     2161 aattctgcag tcgacggtac cgcgggcccg ggatccaccg gatctagata actgatcata
     2221 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     2281 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     2341 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     2401 cattctagtt gtggtttgtc caaactcatc aatgtatctt aacgcgtaaa ttgtaagcgt
     2461 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     2521 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     2581 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     2641 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     2701 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     2761 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     2821 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     2881 taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2941 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     3001 ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg
     3061 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3121 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3181 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3241 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3301 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     3361 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     3421 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     3481 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     3541 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     3601 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     3661 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3721 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     3781 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     3841 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     3901 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     3961 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     4021 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     4081 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     4141 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     4201 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     4261 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     4321 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     4381 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     4441 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     4501 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     4561 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     4621 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     4681 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     4741 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     4801 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     4861 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     4921 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     4981 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     5041 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     5101 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     5161 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     5221 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     5281 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     5341 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     5401 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     5461 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     5521 accgccatgc at
//
