LOCUS       sequence-423482-        6196 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses equimolar Lipid Droplet- and Mitochondria- targeted 
            split-FAST fragments for Short-range LD-MT Contact sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6196)
  AUTHORS   Cali T
  TITLE     SPLICSFAST-P2A
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6196)
  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: 
            "Unpublished"
COMMENT     Sequence Label: SPLICSFAST-LDh-MT-P2A
FEATURES             Location/Qualifiers
     source          1..6196
                     /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      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1165..1221
                     /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             1231..1260
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1787..1804)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1793..2017
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2063..2491
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2150..2169)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2360..2381
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2500..2520)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2505..2834
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2685..2820
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2747..2766
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2901..3695
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2955..2974)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3565..3584
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3869..3990
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3906..3925)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3960..3979
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4039..4055)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4039..4055)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4052..4074)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4063..4079
                     /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(4087..4093,4094..4111,4112..4117))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4132..4153
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4270..4287)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4441..5029)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4521..4540)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5200..5991,5992..6060))
                     /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     5823..5842
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6061..6165)
                     /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 gccaccatgg gagatagcta ctgggtgttc gtgaagcggg aatttggcgg cgatggtggc
      961 agcggcggcg gcgatatcag caagctgatg gacgttctgg tgcccctgct ccagctcctg
     1021 gtcctgctgc tgacactgcc tctgcacctg ctggccctgc tgggctgctg gcagcctctt
     1081 tttgaggcca ttggcaaaat cttcagcaac atcagaatct ctacccagaa agagatcctg
     1141 aaggagctgg catgcggctc tggcgccaca aacttctccc tgctgaaaca ggccggagac
     1201 gtggaagaga atcctggacc aagaagcatg gaacagaagc tgatctctga ggaagatctg
     1261 gaattcgccg ctatggtggg caggaactcc gccatcgccg ctggcgtgtg tggcgctctg
     1321 ttcatcggct actgcatcta tttcgacaga aagagacgga gcgaccccaa cagcggcggc
     1381 ggaggctctg gcggcggagg cagcatggaa cacgtggcct tcggtagcga ggacatcgag
     1441 aacaccctgg ccaagatgga cgacggccag ctggatggac tggcttttgg agccatccag
     1501 ctggacggcg acggcaacat cctgcagtac aacgccgccg agggcgacat cacaggcaga
     1561 gatcctaagc aagtgatcgg caagaacttc ttcaaggacg tggcccctgg caccgacagc
     1621 cctgagttct acggcaagtt caaagaggga gtggccagcg gcaatctgaa caccatgttc
     1681 gagtggatga tccccacctc ccggggccca accaaggtga aggtgcacat gaagaaggcc
     1741 ctgagctgac tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct
     1801 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     1861 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     1921 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     1981 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
     2041 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     2101 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     2161 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     2221 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     2281 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     2341 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     2401 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     2461 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     2521 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     2581 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     2641 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     2701 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     2761 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     2821 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga
     2881 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     2941 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     3001 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     3061 ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg
     3121 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     3181 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     3241 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     3301 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     3361 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     3421 tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     3481 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     3541 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     3601 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     3661 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     3721 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     3781 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     3841 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
     3901 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     3961 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
     4021 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     4081 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     4141 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     4201 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     4261 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     4321 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     4381 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     4441 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     4501 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4561 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4621 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4681 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4741 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4801 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     4861 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     4921 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     4981 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     5041 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     5101 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     5161 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     5221 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     5281 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     5341 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     5401 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     5461 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     5521 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     5581 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     5641 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     5701 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     5761 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     5821 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     5881 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     5941 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     6001 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     6061 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     6121 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     6181 agtgccacct gacgtc
//
