LOCUS       sequence-491010-        6106 bp    DNA     circular SYN 29-OCT-2025
DEFINITION  anchoring mCherry-1x SunTag to the mitochondrial membrane.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6106)
  AUTHORS   Vigano MA, Ell CM, Kustermann MMM, Aguilar G, Matsuda S, Zhao N, 
            Stasevich TJ, Affolter M, Pyrowolakis G
  TITLE     Protein manipulation using single copies of short peptide tags in 
            cultured cells and in Drosophila melanogaster.
  JOURNAL   Development. 2021 Mar 16;148(6):dev191700. doi: 10.1242/dev.191700.
  PUBMED    33593816
REFERENCE   2  (bases 1 to 6106)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 29, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Development."; date: "2021-03-16"; volume: "148(6):dev191700. doi";
            pages: " 10.1242/dev.191700"
COMMENT     Sequence Label: mito_mCherry_SunTag
FEATURES             Location/Qualifiers
     source          1..6106
                     /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"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1212..1916
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1354..1372)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1622..1641
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1989..2045
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     primer_bind     complement(2114..2131)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2120..2344
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2390..2818
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2477..2496)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2687..2708
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2827..2847)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2832..3161
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3012..3147
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3074..3093
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3209..3256
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3275..3649
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    3779..3900
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3816..3835)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3870..3889
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3949..3965)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3949..3965)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3962..3984)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3973..3989
                     /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(3997..4003,4004..4021,4022..4027))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4042..4063
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4180..4197)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4351..4939)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4431..4450)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5110..5901,5902..5970))
                     /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     5733..5752
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5971..6075)
                     /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 ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cgccaccatg agtctgactt
     1021 ccagttccag cgtacgagtt gaatggatcg cagcagttac cattgctgct gggacagctg
     1081 caattggtta tctagcttac aaaagatttt atgttaaaga tcatcgaaat aaagctatga
     1141 taaaccttca catccagaaa gacaacccca agatagtaca tgctggaagc ttaccggtcg
     1201 ccgaattcgg tgtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc
     1261 gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg
     1321 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc
     1381 ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg
     1441 tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt
     1501 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc
     1561 tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg
     1621 gccccgtaat gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg
     1681 aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact
     1741 acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct
     1801 acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac
     1861 agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagggtg
     1921 gtggtggcgc ggccgcagat cgatcgatca cgcgtgccac catgactagt tttgtacaaa
     1981 gcggtccgga agaacttttg agcaagaatt atcatcttga gaacgaagtg gctcgtctta
     2041 agaaaaccgc ggtaaacata tggatcgatc ataatactcg agtctagagg gcccgtttaa
     2101 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     2161 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     2221 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     2281 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     2341 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     2401 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     2461 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     2521 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     2581 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     2641 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     2701 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     2761 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     2821 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2881 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     2941 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3001 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3061 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     3121 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     3181 tatccatttt cggatctgat cagcacgtgt tgacaattaa tcatcggcat agtatatcgg
     3241 catagtataa tacgacaagg tgaggaacta aaccatggcc aagttgacca gtgccgttcc
     3301 ggtgctcacc gcgcgcgacg tcgccggagc ggtcgagttc tggaccgacc ggctcgggtt
     3361 ctcccgggac ttcgtggagg acgacttcgc cggtgtggtc cgggacgacg tgaccctgtt
     3421 catcagcgcg gtccaggacc aggtggtgcc ggacaacacc ctggcctggg tgtgggtgcg
     3481 cggcctggac gagctgtacg ccgagtggtc ggaggtcgtg tccacgaact tccgggacgc
     3541 ctccgggccg gccatgaccg agatcggcga gcagccgtgg gggcgggagt tcgccctgcg
     3601 cgacccggcc ggcaactgcg tgcacttcgt ggccgaggag caggactgac acgtgctacg
     3661 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     3721 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     3781 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     3841 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     3901 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     3961 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     4021 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4081 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4141 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     4201 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     4261 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     4321 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     4381 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     4441 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     4501 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     4561 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     4621 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     4681 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     4741 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     4801 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     4861 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     4921 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     4981 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     5041 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     5101 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     5161 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     5221 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     5281 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     5341 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     5401 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     5461 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     5521 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     5581 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     5641 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     5701 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     5761 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     5821 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     5881 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     5941 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     6001 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     6061 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
