LOCUS       sequence-243601-        7773 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SB-transposon plasmid for stable expression of unphosphorylatable 
            ""dead"" version of AktAR2 variant of FRET-based AKT activity 
            reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7773)
  TITLE     Eric Davis SB transposon plasmids
REFERENCE   2  (bases 1 to 7773)
  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"
COMMENT     Sequence Label: pSBbi-Pur-AktAR2-D
FEATURES             Location/Qualifiers
     source          1..7773
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1130..1150
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1208..1217
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1229..1246
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1250..1282
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(1286..1294,1295..1309)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(1316..1318,1319..1321,1322..2032)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2585..3319
                     /codon_start=1
                     /product="circularly permuted form of the Venus yellow 
                     fluorescent protein (Nagai et al., 2004)"
                     /label=cp173Venus
                     /note="efficient FRET acceptor for CFP"
                     /translation="MGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDH
                     MVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEELFTGVVPILVELDGDVNGHKFSVSG
                     EGEGDATYGKLTLKLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPE
                     GYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHN
                     VYITADKQKNGIKANFKIRHNIE"
     primer_bind     complement(3373..3390)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3379..3603
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        4035..4139
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4140..4208,4209..5000)
                     /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(4358..4377)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5171..5759
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5660..5679
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5913..5930
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6304..6323
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    6322..6377
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     6423..6442
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(6542..7141)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(6625..6645)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     7122..7141
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        join(7773,1..1182)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
ORIGIN
        1 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
       61 gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
      121 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
      181 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg
      241 tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta
      301 cttccacgcc cctggctgca gtacgtgatt cttgatcccg agcttcgggt tggaagtggg
      361 tgggagagtt cgaggccttg cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc
      421 tggcttgggc gctggggccg ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct
      481 gctttcgata agtctctagc catttaaaat ttttgatgac ctgctgcgac gctttttttc
      541 tggcaagata gtcttgtaaa tgcgggccaa gatctgcaca ctggtatttc ggtttttggg
      601 gccgcgggcg gcgacggggc ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg
      661 cgagcgcggc caccgagaat cggacggggg tagtctcaag ctggccggcc tgctctggtg
      721 cctggcctcg cgccgccgtg tatcgccccg ccctgggcgg caaggctggc ccggtcggca
      781 ccagttgcgt gagcggaaag atggccgctt cccggccctg ctgcagggag ctcaaaatgg
      841 aggacgcggc gctcgggaga gcgggcgggt gagtcaccca cacaaaggaa aagggccttt
      901 ccgtcctcag ccgtcgcttc atgtgactcc acggagtacc gggcgccgtc caggcacctc
      961 gattagttct cgagcttttg gagtacgtcg tctttaggtt ggggggaggg gttttatgcg
     1021 atggagtttc cccatactga gtgggtggag actgaagtta ggccagcttg gcacttgatg
     1081 taattctcct tggaatttgc cctttttgag tttggatctt ggttcattct caagcctcag
     1141 acagtggttc aaagtttttt tcttccattt aaggtgtcgt gaaaactacc ccaagctggc
     1201 ctctgaggcc accatggtgc ggggttctca tcatcatcat catcatggta tggctagcat
     1261 gactggtgga cagcaaatgg gtcgggatct gtacgacgat gacgataagg atcccatggt
     1321 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1381 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     1441 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1501 gaccaccctg agctggggcg tgcagtgctt cgcccgctac cccgaccaca tgaagcagca
     1561 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     1621 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     1681 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     1741 ggagtacaac gccatccacg gcaacgtcta tatcaccgcc gacaagcaga agaacggcat
     1801 caaggccaac ttcggcctca actgcaacat cgaggacggc agcgtgcagc tcgccgacca
     1861 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     1921 gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     1981 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agcgcatgca
     2041 taagttttct caagaacaga tcggcgaaaa cattgtgtgc agggtcattt gtaccacggg
     2101 tcaaattccc atccgagatt tgtcagctga tatttcacaa gtgcttaagg aaaaacgatc
     2161 cataaagaaa gtttggacat ttggtagaaa cccagcctgt gactatcatt taggaaacat
     2221 ttcaagactg tcaaataagc atttccaaat actactagga gaagacggta accttttatt
     2281 gaatgacatt tccactaatg ggacctggtt aaatgggcaa aaagtcgaga agaacagcaa
     2341 tcagttactg tctcaaggtg atgaaataac cgttggtgta ggcgtggaat cagatatttt
     2401 atctctggtc attttcataa acgacaaatt taagcagtgc ctcgagcaga acaaagttga
     2461 tcgctctgca ggtaagccag gcagcggcga gggcagcacc aagggcctgg tcgaccctcg
     2521 tccgcgctcg tgcgtgtggc cggaccccag gccggagttt ggaggtaccg gcggcagcga
     2581 gctcatgggc ggcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     2641 ccccgtgctg ctgcccgaca accactacct gagctaccag tccaagctga gcaaagaccc
     2701 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     2761 cggcatggac gagctgtaca agggaggtac cggtggatct atggtgagca agggcgagga
     2821 gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa
     2881 gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagct
     2941 gatctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgggcta
     3001 cggccttcag tgcttcgccc gctaccccga ccacatgaag cagcacgact tcttcaagtc
     3061 cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta
     3121 caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa
     3181 gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa
     3241 cagccacaac gtctatatca ccgccgacaa gcagaagaac ggcatcaagg ccaacttcaa
     3301 gatccgccac aacatcgagt aaggcctgtc aggccaagct tggggatcaa ttctctagag
     3361 ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     3421 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     3481 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     3541 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta
     3601 tggcttctga ggcggaaaga accagctggg cctactagat agcggacccc ttaccgaaac
     3661 atcgccgcat tctgcagagg agtcgaactt agtgtatgta aacttctgac ccactggaat
     3721 tgtgatacag tgaattataa gtgaaataat ctgtctgtaa acaattgttg gaaaaatgac
     3781 ttgtgtcatg cacaaagtag atgtcctaac tgacttgcca aaactattgt ttgttaacaa
     3841 gaaatttgtg gagtagttga aaaacgagtt ttaatgactc caacttaagt gtatgtaaac
     3901 ttccgacttc aactgtatag ggatccaatt ttaacaaaat attaacgctt acaatttcct
     3961 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatca ggtggcactt
     4021 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     4081 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     4141 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     4201 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     4261 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     4321 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     4381 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     4441 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     4501 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     4561 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     4621 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     4681 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     4741 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     4801 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     4861 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     4921 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     4981 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     5041 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     5101 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     5161 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     5221 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     5281 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     5341 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     5401 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     5461 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     5521 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     5581 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     5641 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     5701 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     5761 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     5821 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     5881 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggatc
     5941 cctatacagt tgaagtcgga agtttacata caccttagcc aaatacattt aaactcactt
     6001 tttcacaatt cctgacattt aatcctagta aaaattccct gtcttaggtc agttaggatc
     6061 accactttat tttaagaatg tgaaatatca gaataatagt agagagaatg attcatttca
     6121 gcttttattt ctttcatcac attcccagtg ggtcagaagt ttacatacac tcgactcctc
     6181 tgcagaatgc ggcgatgttt cggtaagggg tcccgagtag ctagttcatg gcagccagca
     6241 tatggcatat gttgccaaac tctaaaccaa atactcattc tgatgtttta aatgatttgc
     6301 cctcccatat gtccttccga gtgagagaca caaaaaattc caacacacta ttgcaatgaa
     6361 aataaatttc ctttattagc cagaagtcag atgctcaagg ggcttcatga tgtccccata
     6421 atttttggca gagggaaaaa gatctcagtg gtatttgtga gccagggcat tggccacacc
     6481 agccaccacc ttctgatagg cagcctgcac ctgaggagtg aattcacgac aggccttcga
     6541 atcaggcacc gggcttgcgg gtcatgcacc aggtgcgcgg tccttcgggc acctcgacgt
     6601 cggcggtgac ggtgaagccg agccgctcgt agaaggggag gttgcggggc gcggaggtct
     6661 ccaggaaggc gggcaccccg gcgcgctcgg ccgcctccac tccggggagc acgacggcgc
     6721 tgcccagacc cttgccctgg tggtcgggcg agacgccgac ggtggccagg aaccacgcgg
     6781 gctccttggg ccggtgcggc gccaggaggc cttccatctg ttgctgcgcg gccagccggg
     6841 aaccgctcaa ctcggccatg cgcgggccga tctcggcgaa caccgccccc gcttcgacgc
     6901 tctccggcgt ggtccagacc gccaccgcgg cgccgtcgtc cgcgacccac accttgccga
     6961 tgtcgagccc gacgcgcgtg aggaagagtt cttgcagctc ggtgacccgc tcgatgtggc
     7021 ggtccggatc gacggtgtgg cgcgtggcgg ggtagtcggc gaacgcggcg gcgagggtgc
     7081 gtacggccct ggggacgtcg tcgcgggtgg cgaggcgcac cgtgggcttg tactcggtca
     7141 tggtggcctc aggtgcagag gtttctacag ggaaagagag aaaggctaca gttagtttgc
     7201 tgtgatgatg ctaagtgccg aggtcagatg tctatggatt gtacacggaa tcccaacgca
     7261 ggaaccaaaa aacactcacc caaaaaaatg gcgctaaggc cgagagaaag gcgaagttcc
     7321 gtctacggct atataccttg aaacaggggc ggcagaagtg acgcaacgcg gttgcgcgga
     7381 cggcttcctg tgttttcatt ggtcgaagcg ttctgcgcgt gcgcagagat tccccaggga
     7441 aggcgggccc tcttccgaat gtccaatccg gacattcttc cggttggacc tccgcggtca
     7501 gcgctgtgcc cggcgagaga tcacgtgggg cgcggaggcg gtgctgctgg ggcacggccg
     7561 tccagcctcg gcggccatat ttttgagggg ctgttcatct cgttcacacg ctctgtccgc
     7621 catgtttgtg agtggaagcg ccattacccc ttcaagcgac tgaaggctgc agggcctctg
     7681 gtggcccgca tggggagacc agacccgcca ggcccgcctt tccgcactca gtccgggctt
     7741 actttatttt gtgagacagg gtctcgccta gag
//
