LOCUS       sequence-457128-        6957 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology repair plasmid for endogenous tagging of AP1muA at the 
            C-terminus with monomeric StayGold. Contains a hygromycin resistance
            cassette for selection of edited cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6957)
  AUTHORS   Stockhammer A, Adarska P, Natalia V, Heuhsen A, Klemt A, Bregu G, 
            Harel S, Rodilla-Ramirez C, Spalt C, Ozsoy E, Leupold P, Grindel A, 
            Fox E, Mejedo JO, Zehtabian A, Ewers H, Puchkov D, Haucke V, 
            Bottanelli F
  TITLE     ARF1 compartments direct cargo flow via maturation into recycling 
            endosomes.
  JOURNAL   Nat Cell Biol. 2024 Oct 4. doi: 10.1038/s41556-024-01518-4.
  PUBMED    39367144
REFERENCE   2  (bases 1 to 6957)
  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: "Nat Cell 
            Biol. 2024 Oct 4. doi: 10.1038/s41556-024-01518-4."
COMMENT     Sequence Label: AP1muA-mStayGold-polyA-Hygro HR
FEATURES             Location/Qualifiers
     source          1..6957
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     742..764
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     761..777
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     761..777
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1857..1978
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(2108..3133)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     enhancer        3286..3477
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     3434..3454
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    3503..3624
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(3751..4428)
                     /codon_start=1
                     /product="bright and highly photostable monomeric green 
                     fluorescent protein (Ando et al., 2023)"
                     /label=mStayGold(J)
                     /note="includes multiple mutations plus an N-terminal 
                     iinker sequence"
                     /translation="MVSTGEELFTGVVPFKFQLKGTINGKSFTVEGEGEGNSHEGSHKG
                     KYVCTSGKLPMSWAALGTSFGYGMKYYTKYPSGLKNWFHEVMPEGFTYDRHIQYKGDGS
                     IHAKHQHFMKNGTYHNIVEFTGQDFKENSPVLTGDMDVSLPNEVQHIPIDDGVECTVTL
                     QYPLLSDESKCVEAYQNTIIKPLHNQPAPDVPFHWIRKQYTQSKDDTEERDHIIQSETL
                     EAHL"
     primer_bind     complement(5622..5639)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5622..5638)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5631..5653)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        5752..5856
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5857..5925,5926..6717)
                     /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     complement(6075..6094)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     protein_bind    6817..6841
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttccagc
      661 gacatatgcc atacgccgaa ctgaactcca cctaatgtct tggagattgt cggatagcgc
      721 ggaataggat tgtgccctct gagcggataa caatttcaca caggaaacag ctatgaccat
      781 gacacatgca tccaccatcg cagacttatc atcactgagc ctccacctag cctgctggtg
      841 acacctcgtg gggccacagg gaaggtcggc gacaaaacca ggcaagccct gtggtcacgc
      901 aggaggagtt gggaccacgc tgcatcagag gaaggctgtg gagtattcaa tgaatggaat
      961 ttaatgtaac tatgacataa acacacacag ggtggggagg acgggtgacc aagcgcagac
     1021 gtgggtgaca cgtggccccg ctctctggac cctcagtggg aaaaagtctg aggctggcgt
     1081 ctctcaccaa accccacctc ccctggtggg tcaatactga tctggctgag tgacagcatc
     1141 tcgtgaccca ggctggccct gggaaggcgc cacaggcgag gcctgcgagt ccagggagca
     1201 ggcagacgct acagtggccc ccgagcgcgc caggtgccag cctcgacgtg taaatggccc
     1261 gctgcgggcg ctgccgggaa ggaagacaag aagggtgagg gctcgtcctc accaagtgct
     1321 tcctgagtgc cgtaggacgt gggtgggaca tggggacagt gagggtaaca caaacagcaa
     1381 cgaatgcaca aagacaagtt ccaggaacac ccataaaccc ataacgttca acaaaatggc
     1441 aacaaaacga cgcccctgct ggcctggcga ggctggcgct tccttcgtgg cagatgccac
     1501 ctgcagctgg ctggctggca agcgttggcc aagctttttg gtacctggga gctcaacagc
     1561 ataaaaggct gtaacctcta cacggaaaaa tttcacatcc ttcccctgaa actgacttca
     1621 ggggcaaaga aacgagactt cttgaagacc agcctcttct gggaaagggg cttctgagac
     1681 gagtgtcgac ggcagagcag atggcctggg gaaggcaggg gtccctgggc agaggctggg
     1741 agggtggcca gagaccaggg agggcccctc catctggtgg gtttggcagg tgtgtcccct
     1801 ggtgctgcca ccaggagctc cgaggctggg gtgttggctg cgacagcccc gaattctaag
     1861 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     1921 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttgg
     1981 ggtgggcgaa gaactccagc atgagatccc cgcgctggag gatcatccag ccggcgtccc
     2041 ggaaaacgat tccgaagccc aacctttcat agaaggcggc ggtggaatcg aaatctcgta
     2101 gtacgtgcta ttcctttgcc ctcggacgag tgctggggcg tcggtttcca ctatcggcga
     2161 gtacttctac acagccatcg gtccagacgg ccgcgcttct gcgggcgatt tgtgtacgcc
     2221 cgacagtccc ggctccggat cggacgattg cgtcgcatcg accctgcgcc caagctgcat
     2281 catcgaaatt gccgtcaacc aagctctgat agagttggtc aagaccaatg cggagcatat
     2341 acgcccggag ccgcggcgat cctgcaagct ccggatgcct ccgctcgaag tagcgcgtct
     2401 gctgctccat acaagccaac cacggcctcc agaagaagat gttggcgacc tcgtattggg
     2461 aatccccgaa catcgcctcg ctccagtcaa tgaccgctgt tatgcggcca ttgtccgtca
     2521 ggacattgtt ggagccgaaa tccgcgtgca cgaggtgccg gacttcgggg cagtcctcgg
     2581 cccaaagcat cagctcatcg agagcctgcg cgacggacgc actgacggtg tcgtccatca
     2641 cagtttgcca gtgatacaca tggggatcag caatcgcgca tatgaaatca cgccatgtag
     2701 tgtattgacc gattccttgc ggtccgaatg ggccgaaccc gctcgtctgg ctaagatcgg
     2761 ccgcagcgat cgcatccatg gcctccgcga ccggctgcag aacagcgggc agttcggttt
     2821 caggcaggtc ttgcaacgtg acaccctgtg cacggcggga gatgcaatag gtcaggctct
     2881 cgctgaactc cccaatgtca agcacttccg gaatcgggag cgcggccgat gcaaagtgcc
     2941 gataaacata acgatctttg tagaaaccat cggcgcagct atttacccgc aggacatatc
     3001 cacgccctcc tacatcgaag ctgaaagcac gagattcttc gccctccgag agctgcatca
     3061 ggtcggagac gctgtcgaac ttttcgatca gaaacttctc gacagacgtc gcggtgagtt
     3121 caggcttttt catatcgatc tttgcaaaag cctaggcctc caaaaaagcc tcctcactac
     3181 ttctggaata gctcagaggc cgaggcggcc tcggcctctg cataaataaa aaaaattagt
     3241 cagccatggg gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct
     3301 gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
     3361 cacctggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct
     3421 ggggactttc cacaccctaa ctgacacaca ttccacagct ggttctttcc gcctcagggc
     3481 ggccgcctta caatttacgc cttaagatac attgatgagt ttggacaaac cacaactaga
     3541 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     3601 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
     3661 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtatggct
     3721 gattatgatc tagagtcgcg ggctagctta caggtgggcc tccagggtct cgctctggat
     3781 gatgtggtct ctctcctcgg tgtcgtcctt gctctgggtg tactgctttc tgatccagtg
     3841 gaatggcaca tcgggggctg gctgattgtg caggggcttg atgatggtgt tctggtaggc
     3901 ctccacgcac ttgctctcgt cgctcagcag agggtactgc agggtcactg tgcactccac
     3961 gccgtcatct atggggatgt gctgcacctc gttgggcagg ctcacgtcca tgtcgccggt
     4021 cagcacgggg ctgttctcct tgaagtcctg gccggtgaac tccacgatgt tgtggtaggt
     4081 gccgttcttc atgaagtgct ggtgcttggc gtggatgctg ccgtcgccct tgtactggat
     4141 gtgtctgtcg taggtgaagc cctcgggcat cacctcgtgg aaccagttct tcaggccgct
     4201 ggggtacttg gtgtagtact tcatgccata gccgaagcta gttcccaggg cggcccaaga
     4261 cattggcagt ttgccgctgg tgcacacgta cttgcctttg tggctgccct cgtggctatt
     4321 gccctcgccc tcgccttcca cggtgaagct cttgccgttg atggtgccct tcagctggaa
     4381 cttgaagggc accacgccgg taaacagctc ctcgcctgta gacaccatgg atcctgagcc
     4441 ggaaccagag cctgaccctg atccctgggt ccggagctgg taatctagga gacaggaggg
     4501 aggcgtcatg ctgggctctg caccctccct gctttcggga cccgacccga tggctccacc
     4561 ctgggggcag ccacagagga cccctcagta caaccccacc ccacgggtag ggcctcactc
     4621 acctccattc tgcgtgatat aacgcaccca gggcagggcc tggtacccac tcttctcaat
     4681 gatcttcagg tagcgcacct ggggcacagg tagcagggcc ctgagcaccg caggctcccg
     4741 ccctgttggt ccccccttaa tctttcctgc tggggcagcc tggggctggc aggggaggcc
     4801 atgacgacag gatgagcaca ggcctcaggg gggtgaaagg gatttgtctg gaccatgcaa
     4861 atgtgtggga gggcagagcc gcctctggaa gtggtgagca gccaggatgt ggcttgcaga
     4921 cacggcctca gcagcctgct tggggggccc ggccagaagg cccctggctc tctgggagcc
     4981 tctcctccct ctgtgggctt ttgagggggt gccagggaaa agaggtccca tggggcatga
     5041 gtggacggca gagcctcaaa tgctagtttg gggcagaatg agtgagcttt gagctcaggg
     5101 gtcagggctg agcagaggca gcctggcctc accctgtctg tggggaggca gaggcccaca
     5161 gcacagcagg gtcctgattg cagctgactg acaccctttc ccaagctggc tgcattgata
     5221 tctgtcacct gcggcctctg tccctgttct gggccccggc cgcctgggcc ttacgtacct
     5281 ggatgccgga ggtagtgaag taagggatct cgaacttgac actgatcggg ggcttgccct
     5341 ccttgtcttc ggcctccaca ctaggcaggc cgaagtgggc ccgcatcagg tactccttgc
     5401 cgccctgggg agcagacggg ggaggcgctc aggcctaggc cctgccccag cttagccaac
     5461 agctctggcc ctagtggcga tgggcagaga ctgaccactc accccagtcc agggagggga
     5521 gcatgagccc ctgtgctggg gagccaaagc tcggccctgt gctgccccaa ggtcaggcgg
     5581 aatggcactt ccttattcca gatgcgtgcg gattatgcat gactggccgt cgttttacaa
     5641 cgtcgtgact gggaaaaccc tggcgcttgt ggtggatcac agttattgga gacttccttg
     5701 gccgtacgat gggttttgtt tcaggtatct ggcacttttc ggggaaatgt gcgcggaacc
     5761 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     5821 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     5881 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     5941 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     6001 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     6061 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     6121 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     6181 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     6241 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     6301 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     6361 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     6421 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     6481 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     6541 attgctgata aatctggagc cggtgagcgt ggctctcgcg gtatcattgc agcactgggg
     6601 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     6661 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     6721 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     6781 aggatctagg tgaagatcct tgaattgcca ttatcaacaa gtttgtacaa aaaagcaggc
     6841 tccgaattgg tatcacgagg acgatttgaa gccccctacc catttacgct gacttgacgg
     6901 gacggcggct ttgttgaata gagcgtcaga ccccgtagaa aagatcaaag gatcttc
//
