LOCUS       sequence-360359-        6065 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of SNAP-Tagged Cox8A in Mammalian cells .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6065)
  AUTHORS   Ana Egana
  TITLE     Egana Lab NEB plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6065)
  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            "Unpublished"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: pSNAPf-Cox8A Control Plasmid
FEATURES             Location/Qualifiers
     source          1..6065
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             696..770
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             951..1496
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     intron          1607..1836
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     1819..1838
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    1909..2482
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2076..2093)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2303..2322
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2518..3321
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2581..2600)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3191..3210
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3451..3572
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3488..3507)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3674..3690)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3674..3690)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3687..3709)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3698..3714
                     /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(3722..3728,3729..3746,3747..3752))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3767..3788
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3905..3922)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4076..4664)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4156..4175)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4835..5626,5627..5695))
                     /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     5458..5477
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5696..5800)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5875..5894)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccgagc tcggatcgat ggccgcaagc ttaccatgtc cgtcctgacg ccgctgctgc
      721 tgcggggctt gacaggctcg gcccggcggc tcccagtgcc gcgcgccaag atccattcgt
      781 tgccgccgga ggggaagctt gggatcatgg aattggccgt tgggcttacc tcctgcttcg
      841 tgaccttcct cctgccagcg ggctggatcc tgtcacacct ggagacctac aggaggccag
      901 aggatatcgg cgcgccagca tttaaatctg tacagaccgg tgaattcacc atggacaaag
      961 actgcgaaat gaagcgcacc accctggata gccctctggg caagctggaa ctgtctgggt
     1021 gcgaacaggg cctgcaccgt atcatcttcc tgggcaaagg aacatctgcc gccgacgccg
     1081 tggaagtgcc tgccccagcc gccgtgctgg gcggaccaga gccactgatg caggccaccg
     1141 cctggctcaa cgcctacttt caccagcctg aggccatcga ggagttccct gtgccagccc
     1201 tgcaccaccc agtgttccag caggagagct ttacccgcca ggtgctgtgg aaactgctga
     1261 aagtggtgaa gttcggagag gtcatcagct acagccacct ggccgccctg gccggcaatc
     1321 ccgccgccac cgccgccgtg aaaaccgccc tgagcggaaa tcccgtgccc attctgatcc
     1381 cctgccaccg ggtggtgcag ggcgacctgg acgtgggggg ctacgagggc gggctcgccg
     1441 tgaaagagtg gctgctggcc cacgagggcc acagactggg caagcctggg ctgggtcctg
     1501 caggcggatc cgcgtttaaa ctcgaggtta attaatgagc ggccgcatag ataactgatc
     1561 cagtgtgctg gaattaattc gctgtctgcg agggccagct gttggggtga gtactccctc
     1621 tcaaaagcgg gcatgacttc tgcgctaaga ttgtcagttt ccaaaaacga ggaggatttg
     1681 atattcacct ggcccgcggt gatgcctttg agggtggccg cgtccatctg gtcagaaaag
     1741 acaatctttt tgttgtcaag cttgaggtgt ggcaggcttg agatctggcc atacacttga
     1801 gtgacaatga catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
     1861 aggtcgagca tgcatctagg gcggccaatt ccgcccctct cccccccccc cttttccctc
     1921 ccccccccct aacgttactg gccgaagccg cttggaataa ggccggtgtg cgtttgtcta
     1981 tatgttattt tccaccatat tgccgtcttt tggcaatgtg agggcccgga aacctggccc
     2041 tgtcttcttg acgagcattc ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct
     2101 gttgaatgtc gtgaaggaag cagttcctct ggaagcttct tgaagacaaa caacgtctgt
     2161 agcgaccctt tgcaggcagc ggaacccccc acctggcgac aggtgcctct gcggccaaaa
     2221 gccacgtgta taagatacac ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg
     2281 gatagttgtg gaaagagtca aatggctctc ctcaagcgta ttcaacaagg ggctgaagga
     2341 tgcccagaag gtaccccatt gtatgggatc tgatctgggg cctcggtgca catgctttac
     2401 atgtgtttag tcgaggttaa aaaaacgtct aggccccccg aaccacgggg acgtggtttt
     2461 cctttgaaaa acacgatgat aagcttgcca caacccggga taattcctgc agccaatatg
     2521 ggatcggcca ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     2581 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     2641 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     2701 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     2761 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     2821 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     2881 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     2941 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     3001 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg
     3061 cccgacggcg atgatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     3121 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     3181 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     3241 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     3301 cttcttgacg agttcttctg aggggatcaa ttctctagat aactgatcat aatcagccat
     3361 accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg
     3421 aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac
     3481 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     3541 tgtggtttgt ccaaactcat caatgtatct taacgcgtcg agtgcattct agttgtggtt
     3601 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     3661 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     3721 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     3781 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     3841 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     3901 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     3961 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4021 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4081 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4141 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4201 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4261 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4321 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4381 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4441 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     4501 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4561 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     4621 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     4681 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     4741 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     4801 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     4861 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     4921 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     4981 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5041 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5101 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     5161 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5221 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     5281 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     5341 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     5401 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     5461 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     5521 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     5581 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     5641 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     5701 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     5761 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     5821 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat
     5881 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc
     5941 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat
     6001 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
     6061 gcgtt
//
