LOCUS       sequence-429346-        4194 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Endogenously tag human TMEM115 with a C-terminal 3xHA tag in 
            addition to cytosolic mScarlet-I for cell sorting..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4194)
  AUTHORS   Hollingsworth LR, Veeraraghavan P, Paulo JA, Harper JW
  TITLE     Spatiotemporal proteomic profiling of cellular responses to NLRP3 
            agonists
  JOURNAL   bioRxiv 2024.04.19.590338
REFERENCE   2  (bases 1 to 4194)
  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: "bioRxiv 
            2024.04.19.590338"
COMMENT     Sequence Label: hTMEM115-TEV-3xHA-P2A-mScarlet HDR template
FEATURES             Location/Qualifiers
     source          1..4194
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             17..73
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             74..769
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(207..225)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     475..494
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(1284..1300)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1284..1300)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1297..1319)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1472..2060)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1552..1571)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2177..2201)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2301..3092,3093..3161))
                     /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     2924..2943
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3162..3266)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3365..3387
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3379..3396
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3380..3396
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4094..4114
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             4115..4141
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4145..4171
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(4175..4194,1..7)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
ORIGIN
        1 ctacgcagga tccggcgcaa caaacttctc tctgctgaaa caagccggag atgtcgaaga
       61 gaatcctgga ccgatggtga gcaagggcga ggcagtgatc aaggagttca tgcggttcaa
      121 ggtgcacatg gagggctcca tgaacggcca cgagttcgag atcgagggcg agggcgaggg
      181 ccgcccctac gagggcaccc agaccgccaa gctgaaggtg accaagggtg gccccctgcc
      241 cttctcctgg gacatcctgt cccctcagtt catgtacggc tccagggcct tcatcaagca
      301 ccccgccgac atccccgact actataagca gtccttcccc gagggcttca agtgggagcg
      361 cgtgatgaac ttcgaggacg gcggcgccgt gaccgtgacc caggacacct ccctggagga
      421 cggcaccctg atctacaagg tgaagctccg cggcaccaac ttccctcctg acggccccgt
      481 aatgcagaag aagacaatgg gctgggaagc gtccaccgag cggttgtacc ccgaggacgg
      541 cgtgctgaag ggcgacatta agatggccct gcgcctgaag gacggcggcc gctacctggc
      601 ggacttcaag accacctaca aggccaagaa gcccgtgcag atgcccggcg cctacaacgt
      661 cgaccgcaag ttggacatca cctcccacaa cgaggactac accgtggtgg aacagtacga
      721 acgctccgag ggccgccact ccaccggcgg catggacgag ctgtacaagt aactccagac
      781 caccttgagt gtggcacctc ccctcccaag ccccccttga catcctctca gctactccag
      841 ggcacctgac tgctctgagg agagggaaga aggcctgctg gggctttcca tggccttctg
      901 ctgtttctcg ccaacactac ccaggactct tgctacctgg ttccaactcc agacaaccac
      961 tatgccaggc ccggagcctc tgaggcatcg gccagtccag gccctcatct gaggtaagaa
     1021 tgtacatcag ctggcagccc caagcaagtg gctgcaggga cactgatgcc acagctcctg
     1081 ggccggccct cacatctgaa actggttgcc gagagccctg agccaaggca aggatttgcc
     1141 aaaaatgttc tgggggccca gcaaatgcag gagccgacct ggggctgcac atccctgccc
     1201 atccccagaa agactgttcc tgtcaggagg ctaggtggag gctcagtgat gataagtctg
     1261 cgatggtgga tgcatgtgtc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     1321 agagggcaca atcctattcc gcgctatccg acaatctcca agacattagg tggagttcag
     1381 ttcggcgtat ggcatatgtc gctggaaaga acatgtgagc aaaaggccag caaaaggcca
     1441 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     1501 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     1561 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     1621 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     1681 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     1741 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     1801 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     1861 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     1921 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     1981 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     2041 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctctat
     2101 tcaacaaagc cgccgtcccg tcaagtcagc gtaaatgggt agggggcttc aaatcgtcct
     2161 cgtgatacca attcggagcc tgcttttttg tacaaacttg ttgataatgg caattcaagg
     2221 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     2281 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     2341 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     2401 gagggcttac catctggccc cagtgctgca atgataccgc gagagccacg ctcaccggct
     2461 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     2521 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     2581 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     2641 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     2701 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     2761 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     2821 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     2881 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     2941 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     3001 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     3061 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     3121 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     3181 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     3241 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccaga tacctgaaac
     3301 aaaacccatc gtacggccaa ggaagtctcc aataactgtg atccaccaca agcgccaggg
     3361 ttttcccagt cacgacgttg taaaacgacg gccagtcatg cataatccgc acgcatctgg
     3421 aataaggaag tgccattccg cctgacctca ggagaggggg agaattaata acccttccct
     3481 tggatgtgat gccagcaccc aggcagaaac tcctggaaac tccagaactt gccgttccac
     3541 ccatctgccc tagggcacta gttccactga cccttagata ccaccctcag aaccattcct
     3601 ggtggctggc ctcgtgggca gtctggcaga cacccagctg accttcccat cccggaggtg
     3661 cctcagctat ggagggaaga ttcttagagc ctttgggtta tcattctgat ggcagagtgg
     3721 gtcctaaacc aatctctggg gagtcccagg ctctgcatgg tgaaggggaa acataggatt
     3781 ctgggggagg agtctaggtg gtacggtgag gagtcagcct gctgcggaag ggtagagggc
     3841 atgagccaag gccaaaccca gagtgacctt tcctctctcc tctggcccca ggcaactggc
     3901 actcaaagcc ctcaatgagc ggctgaagag agtggaagac cagtccatct ggcccagcat
     3961 ggatgatgat gaagaggagt ctggggccaa ggtggacagc cccctgccct cagacaaagc
     4021 tcccacaccc ccagggaagg gggctgcccc agaatccagt ctaatcacct tcgaggcagc
     4081 tcccccgacg ctggaaaacc tgtacttcca gggctacccg tatgatgttc cggattacgc
     4141 tggctatccc tacgacgtgc ccgactatgc cgggtacccc tatgacgtcc caga
//
