LOCUS       Exported                7368 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  cre-dependent CA-AMPK under EF1a promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-EF1a-Flex-CA-AMPK-H150R-T2A-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7368)
  AUTHORS   Kong D, Dagon Y, Campbell JN, Guo Y, Yang Z, Yi X, Aryal P, 
            Wellenstein K, Kahn BB, Sabatini BL, Lowell BB
  TITLE     A Postsynaptic AMPK-->p21-Activated Kinase Pathway Drives 
            Fasting-Induced Synaptic Plasticity in AgRP Neurons.
  JOURNAL   Neuron. 2016 Jun 16. pii: S0896-6273(16)30204-5. doi: 
            10.1016/j.neuron.2016.05.025.
  PUBMED    27321921
REFERENCE   2  (bases 1 to 7368)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7368
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    1450..1483
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    1534..1567
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     CDS             complement(1576..2283)
                     /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(1854..1873)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2123..2141
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2290..2346)
                     /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"
     protein_bind    complement(3353..3386)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    complement(3437..3470)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    3495..4083
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3548..3568)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3966..3977)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4115..4591
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4314..4333)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4631..4771
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4631..4760
                     /label=AAV2 ITR
     rep_origin      4846..5301
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4933..4952)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5143..5164
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5318..5337)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5437..5459
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5497..5515)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5583..5687
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5688..6548
                     /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(5906..5925)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6719..7307
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7208..7227
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc
     1441 gactccggaa taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga
     1501 ttgtagctgc tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg
     1561 aagttatggc gcgccttact tgtacagctc gtccatgccg ccggtggagt ggcggccctc
     1621 ggcgcgttcg tactgttcca cgatggtgta gtcctcgttg tgggaggtga tgtccaactt
     1681 gatgttgacg ttgtaggcgc cgggcagctg cacgggcttc ttggccttgt aggtggtctt
     1741 gacctcagcg tcgtagtggc cgccgtcctt cagcttcagc ctctgcttga tctcgccctt
     1801 cagggcgccg tcctcggggt acatccgctc ggaggaggcc tcccagccca tggtcttctt
     1861 ctgcattacg gggccgtcgg aggggaagtt ggtgccgcgc agcttcacct tgtagatgaa
     1921 ctcgccgtcc tgcagggagg agtcctgggt cacggtcacc acgccgccgt cctcgaagtt
     1981 catcacgcgc tcccacttga agccctcggg gaaggacagc ttcaagtagt cggggatgtc
     2041 ggcggggtgc ttcacgtagg ccttggagcc gtacatgaac tgaggggaca ggatgtccca
     2101 ggcgaagggc agggggccac ccttggtcac cttcagcttg gcggtctggg tgccctcgta
     2161 ggggcggccc tcgccctcgc cctcgatctc gaactcgtgg ccgttcacgg agccctccat
     2221 gtgcaccttg aagcgcatga actccttgat gatggccatg ttatcctcct cgcccttgct
     2281 cacctcgaga ggtccagggt tctcctccac gtctccagcc tgcttcagca ggctgaagtt
     2341 agtagctccg cttccgggct tcttctctcc acccgtgagc accagagcct gtaagatgtc
     2401 agacagcgaa acgatgccct tgaccacgtc gtgttcatcc accaccacca gacggtgaac
     2461 ctctgcctcc accagcctat tgatgatggt ttccagagtc tcatgcaggt agcatttgag
     2521 aacaccctca aagtagtggg accgatgctg cagggctttt gtcacagaca catctaggtt
     2581 gttgtaggtc ttttcggctg ccaaattgat cacatcaaac ttggagtaga tgtccaccac
     2641 acgccctttc tcatccacta caggtaaggc ggagactcgg tgctgtacaa agatgcccag
     2701 agccacgtag acaggcgtgg tagtacggac catggcaata ttggcatagg tgccaatctg
     2761 cagctcttgg agagacttag acatgaactc cggcttgggg aactcggtga taaacaactt
     2821 gaggaacttg aggatccgct tgtgagtaag gatgtacaag gtgttgcctg actctgggtc
     2881 gataactggg agcctgcgga tcttatttcg aattaatgaa gagacagcat caaacaagct
     2941 ggcatttgga gagatgcaga caagtggctt aaaggagtcc tgcaggtaca cctctctcca
     3001 cgtctctatc ttgtgctcct ccagttcgta gatctgcacc agggctgact tatagtatcg
     3061 gtgcaaaatg ttgatgaagt cggtgatggt cagcatgccc acaaaactct gcttcttact
     3121 gtcccacaaa ggggcggcac gaacaccatt ggtcaccagg gcaaaaaagg ctttctttac
     3181 ctgtagcgaa gtgtcaaata ccaccaactt ggaacttgtg ggaattaggt catagcagcg
     3241 atgagacttc atgaaggaag tatacacact attgtttgat tccggggtct cttgaaagtg
     3301 ttcattctct agagctggcg agctctctgc agcaaccgac tccattgcta gcataacttc
     3361 gtataaagta tcctatacga agttatttgc cttaacccag aaattatcac tgttattctt
     3421 tagaatggtg caaagaataa cttcgtataa tgtatgctat acgaagttat gaattcgata
     3481 tcaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc
     3541 ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg
     3601 ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc
     3661 tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg
     3721 acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg
     3781 ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga
     3841 caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct
     3901 ttccttggct gctcgcctat gttgccacct ggattctgcg cgggacgtcc ttctgctacg
     3961 tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc
     4021 ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc
     4081 cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg catccctgtg acccctcccc
     4141 agtgcctctc ctggccctgg aagttgccac tccagtgccc accagccttg tcctaataaa
     4201 attaagttgc atcattttgt ctgactaggt gtccttctat aatattatgg ggtggagggg
     4261 ggtggtatgg agcaaggggc aagttgggaa gacaacctgt agggcctgcg gggtctattg
     4321 ggaaccaagc tggagtgcag tggcacaatc ttggctcact gcaatctccg cctcctgggt
     4381 tcaagcgatt ctcctgcctc agcctcccga gttgttggga ttccaggcat gcatgaccag
     4441 gctcagctaa tttttgtttt tttggtagag acggggtttc accatattgg ccaggctggt
     4501 ctccaactcc taatctcagg tgatctaccc accttggcct cccaaattgc tgggattaca
     4561 ggcgtgaacc actgctccct tccctgtcct tctgattttg taggtaacca cgtgcggacc
     4621 gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg
     4681 ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca
     4741 gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt tctccttacg
     4801 catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg
     4861 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     4921 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     4981 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     5041 tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga
     5101 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     5161 ctggaacaac actcaaccct atctcgggct attcttttga tttataaggg attttgccga
     5221 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     5281 aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat
     5341 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     5401 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     5461 tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat
     5521 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg
     5581 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     5641 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     5701 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5761 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     5821 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     5881 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     5941 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     6001 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     6061 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     6121 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     6181 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     6241 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     6301 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     6361 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     6421 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     6481 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     6541 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     6601 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     6661 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     6721 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     6781 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     6841 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     6901 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     6961 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     7021 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     7081 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     7141 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     7201 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     7261 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     7321 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgt
//
