LOCUS       sequence-415625-        5486 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses outer mitochondria-targeted split-NFAST for Short-range 
            ER-Mito Contact sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5486)
  TITLE     Mammalian expression plasmids for ER-Mitochondria contact sites 
            monitoring using splitFAST (OMM-SPLICS-NFAST and ER-SPLICS-CFAST10)
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5486)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: OMM-SPLICSs-nFAST
FEATURES             Location/Qualifiers
     source          1..5486
                     /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"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      919..928
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             928..957
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      964..973
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1494..1511)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1500..1724
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1770..2198
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1857..1876)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2067..2088
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2207..2227)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2212..2541
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2392..2527
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2454..2473
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        2589..2636
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2655..3029
                     /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    3159..3280
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3196..3215)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3250..3269
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3329..3345)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3329..3345)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3342..3364)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3353..3369
                     /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(3377..3383,3384..3401,3402..3407))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3422..3443
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3560..3577)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3731..4319)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3811..3830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4490..5281,5282..5350))
                     /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     5113..5132
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5351..5455)
                     /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 ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gctaccggac tcagatctgc caccatggaa caaaagctta tttctgaaga ggacttggaa
      961 ttcgccgcca tggtgggccg gaacagcgcc atcgccgcgg gcgtgtgcgg tgccctcttc
     1021 atagggtact gcatctactt tgaccgcaaa aggcggagtg accccaactc cggaggaggc
     1081 ggcagcggcg gagggggatc catggagcat gttgcctttg gcagtgagga catcgagaac
     1141 actctggcca aaatggacga cggacaactg gatgggttgg cctttggcgc aattcagctc
     1201 gatggtgacg ggaatatcct gcagtacaat gctgctgaag gagacatcac aggcagagat
     1261 cccaaacagg tgattgggaa gaacttcttc aaggatgttg cacctggaac ggattctccc
     1321 gagttttacg gcaaattcaa ggaaggcgta gcgtcaggga atctgaacac catgttcgaa
     1381 tggatgatac cgacaagcag gggaccaacc aaggtcaagg tgcacatgaa gaaagccctt
     1441 tcctaatagg cggccgctcg agtctagagg gcccgtttaa acccgctgat cagcctcgac
     1501 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     1561 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     1621 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     1681 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     1741 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     1801 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     1861 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     1921 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     1981 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     2041 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     2101 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     2161 aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag
     2221 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2281 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2341 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     2401 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     2461 gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt
     2521 ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat
     2581 cagcacgtgt tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg
     2641 tgaggaacta aaccatggcc aagttgacca gtgccgttcc ggtgctcacc gcgcgcgacg
     2701 tcgccggagc ggtcgagttc tggaccgacc ggctcgggtt ctcccgggac ttcgtggagg
     2761 acgacttcgc cggtgtggtc cgggacgacg tgaccctgtt catcagcgcg gtccaggacc
     2821 aggtggtgcc ggacaacacc ctggcctggg tgtgggtgcg cggcctggac gagctgtacg
     2881 ccgagtggtc ggaggtcgtg tccacgaact tccgggacgc ctccgggccg gccatgaccg
     2941 agatcggcga gcagccgtgg gggcgggagt tcgccctgcg cgacccggcc ggcaactgcg
     3001 tgcacttcgt ggccgaggag caggactgac acgtgctacg agatttcgat tccaccgccg
     3061 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     3121 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     3181 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     3241 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     3301 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     3361 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     3421 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     3481 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3541 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3601 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3661 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3721 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3781 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3841 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3901 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3961 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4021 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4081 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4141 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     4201 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4261 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4321 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4381 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4441 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4501 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4561 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4621 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4681 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4741 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4801 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4861 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4921 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4981 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     5041 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     5101 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     5161 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     5221 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     5281 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     5341 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     5401 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     5461 ttccccgaaa agtgccacct gacgtc
//
