LOCUS       sequence-106989-        7649 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty AAV-RAM plasmid w. multiple cloning site (MCS).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7649)
  AUTHORS   Sorensen AT, Cooper YA, Baratta MV, Weng FJ, Zhang Y, Ramamoorthi K,
            Fropf R, LaVerriere E, Xue J, Young A, Schneider C, Gotzsche CR, 
            Hemberg M, Yin JC, Maier SF, Lin Y
  TITLE     A robust activity marking system for exploring active neuronal 
            ensembles.
  JOURNAL   Elife. 2016 Sep 23;5. pii: e13918. doi: 10.7554/eLife.13918.
  PUBMED    27661450
REFERENCE   2  (bases 1 to 7649)
  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: "Elife."; 
            date: "2016-09-23"; volume: "5. pii: e13918. doi"; pages: " 
            10.7554/eLife.13918"
COMMENT     Sequence Label: pAAV-RAM-d2TTA::TRE-MCS-WPRE-pA
FEATURES             Location/Qualifiers
     source          1..7649
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      2..457
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     299..320
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        484..588
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(589..657,658..1449)
                     /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(807..826)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1620..2208
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2109..2128
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     mobile_element  2511..2735
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2802..3335)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3524..3529,3530..3546,3547..3552))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     repeat_region   3922..4051
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        4065..4379
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    4073..4091
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4109..4127
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4144..4162
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4180..4198
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4216..4234
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4251..4269
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4287..4305
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     4352..4376
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     misc_feature    4446..5034
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4499..4519)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4917..4928)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5070..5087)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5076..5187
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     polyA_signal    5335..5383
                     /note="synthetic polyadenylation signal"
     misc_feature    5397..5488
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     5437..5456
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     CDS             join(5854..6471,6472..6477,6478..6513,6514..6516,6517..6552,
                     6553..6555,6556..6591,6592..6600)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=tTA-Advanced
                     /note="In the Tet-Off(R) system, tTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     primer_bind     complement(6613..6629)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6654..6673)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6655..6673)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    6699..6833
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     repeat_region   6863..7003
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6863..6992
                     /label=AAV2 ITR
     polyA_signal    7140..7274
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     mobile_element  7303..7468
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
      301 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
      361 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
      421 aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt tcggggaaat
      481 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
      541 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
      601 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
      661 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
      721 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
      781 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
      841 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
      901 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
      961 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     1021 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     1081 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     1141 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     1201 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     1261 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     1321 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     1381 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     1441 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     1501 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     1561 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     1621 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     1681 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     1741 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     1801 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     1861 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     1921 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     1981 tacaccgaac tgagatacct acagcgtgag cattgagaaa gcgccacgct tcccgaaggg
     2041 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     2101 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     2161 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     2221 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     2281 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     2341 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     2401 cggtattttc tccttacgca tctgtgcggt atttcacacc gcagaccagc cgcgtaacct
     2461 ggcaaaatcg gttacggttg agtaataaat ggatgccctg cgtaagcggg tgtgggcgga
     2521 caataaagtc ttaaactgaa caaaatagat ctaaactatg acaataaagt cttaaactag
     2581 acagaatagt tgtaaactga aatcagtcca gttatgctgt gaaaaagcat actggacttt
     2641 tgttatggct aaagcaaact cttcattttc tgaagtgcaa attgcccgtc gtattaaaga
     2701 ggggcgtggc caagggcatg gtaaagacta tattcgcggc gttgtgacaa tttaccgaac
     2761 aactccgcgg ccgggaagcc gatctcggct tgaacgaatt gttaggtggc ggtacttggg
     2821 tcgatatcaa agtgcatcac ttcttcccgt atgcccaact ttgtatagag agccactgcg
     2881 ggatcgtcac cgtaatctgc ttgcacgtag atcacataag caccaagcgc gttggcctca
     2941 tgcttgagga gattgatgag cgcggtggca atgccctgcc tccggtgctc gccggagact
     3001 gcgagatcat agatatagat ctcactacgc ggctgctcaa acctgggcag aacgtaagcc
     3061 gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta
     3121 cggagcaagt tcccgaggta atcggagtcc ggctgatgtt gggagtaggt ggctacgtct
     3181 ccgaactcac gaccgaaaag atcaagagca gcccgcatgg atttgacttg gtcagggccg
     3241 agcctacatg tgcgaatgat gcccatactt gagccaccta actttgtttt agggcgactg
     3301 ccctgctgcg taacatcgtt gctgctgcgt aacatcgttg ctgctccata acatcaaaca
     3361 tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga ggcatagact gtacaaaaaa
     3421 acagtcataa caagccatga aaaccgccac tgcgccgtta ccaccgctgc gttcggtcaa
     3481 ggttctggac cagttgcgtg agcgcatacg ctacttgcat tacagtttac gaaccgaaca
     3541 ggcttatgtc aactgggttc gtgccttcat ccgtttccac ggtgtgcgtc acccggcaac
     3601 cttgggcagc agcgaagtcg aggcatttct gtcctggctg gcgaacgagc gcaaggtttc
     3661 ggtctccacg catcgtcagg cattggcggc cttgctgttc ttctacggca aggtgctgtg
     3721 cacggatctg ccctggcttc aggagatcgg aagacctcgg ccgtcgcggc gcttgccggt
     3781 ggtgctgacc ccggatgaag tggttcgcat cctcggtttt ctggaaggcg agcatcgttt
     3841 gttcgcccag gactctagct atagttctag tggttggcta cattattgaa gcatttatca
     3901 gggttattgt ctcagagcat gcctgcaggc agctgcgcgc tcgctcgctc actgaggccg
     3961 cccgggcgtc gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag
     4021 ggagtggcca actccatcac taggggttcc tgcggccgca tctcgagttt actccctatc
     4081 agtgatagag aacgtatgtc gagtttactc cctatcagtg atagagaacg atgtcgagtt
     4141 tactccctat cagtgataga gaacgtatgt cgagtttact ccctatcagt gatagagaac
     4201 gtatgtcgag tttactccct atcagtgata gagaacgtat gtcgagttta tccctatcag
     4261 tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgta tgtcgaggta
     4321 ggcgtgtacg gtgggaggcc tatataagca gagctcgttt agtgaaccgt cagatcgcca
     4381 ccggtgttta aacgctagcg gcgcgccagc gatccaccgg atctagataa ctgatcctta
     4441 tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     4501 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     4561 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     4621 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     4681 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     4741 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     4801 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     4861 tgctcgcctg tgttgccacc tggattctgc gcgggacgcc cttctgctac gtcccttcgg
     4921 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
     4981 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcctgatc
     5041 gataccgtcg actagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     5101 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     5161 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     5221 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca gactagtacg
     5281 tctctcaagg ataagtaagt aatattaagg tacgggaggt attggacagg ccgcaataaa
     5341 atatctttat tttcattaca tctgtgtgtt ggttttttgt gtgaatcgat agtactaaca
     5401 tacgctctcc atcaaaacaa aacgaaacaa aacaaactag caaaataggc tgtccccagt
     5461 gcaagtgcag gtgccagaac atttctctgg cctaactggc cgacgcgtac cctagaagtt
     5521 tgttcgtgac tcagactaga agtttgttcg tgactcagac tagaagtttg ttcgtgactc
     5581 agactagaag tttgttcgtg actcagaact cattcataaa acgcttgtta taaaagcagt
     5641 ggctgcggcg cctcgtactc caaccgcatc tgcagcgagc aactgagaag ccaagactga
     5701 gccggcggcc ggatccagct catgagccat ggcttcccgc cggaggtgga ggagcaggat
     5761 gatggcacgc tgcccatgtc ttgtgcccag gagagcggga tggaccgtca ccctgcagcc
     5821 tgtgcttctg ctaggatcaa tgtggaattc accatgtcta gactggacaa gagcaaagtc
     5881 ataaactctg ctctggaatt actcaatgaa gtcggtatcg aaggcctgac gacaaggaaa
     5941 ctcgctcaaa agctgggagt tgagcagcct accctgtact ggcacgtgaa gaacaagcgg
     6001 gccctgctcg atgccctggc aatcgagatg ctggacaggc atcataccca cttctgcccc
     6061 ctggaaggcg agtcatggca agactttctg cggaacaacg ccaagtcatt ccgctgtgct
     6121 ctcctctcac atcgcgacgg ggctaaagtg catctcggca cccgcccaac agagaaacag
     6181 tacgaaaccc tggaaaatca gctcgcgttc ctgtgtcagc aaggcttctc cctggagaac
     6241 gcactgtacg ctctgtccgc cgtgggccac tttacactgg gctgcgtatt ggaggatcag
     6301 gagcatcaag tagcaaaaga ggaaagagag acacctacca ccgattctat gcccccactt
     6361 ctgagacaag caattgagct gttcgaccat cagggagccg aacctgcctt ccttttcggc
     6421 ctggaactaa tcatatgtgg cctggagaaa cagctaaagt gcgaaagcgg cgggccggcc
     6481 gacgcccttg acgattttga cttagacatg ctcccagccg atgcccttga cgactttgac
     6541 cttgatatgc tgcctgctga cgctcttgac gattttgacc ttgacatgct ccccgggtaa
     6601 ctaagtaagc atcgataccg tcgacctcga gggggggccc ggtacccaat tcgccctata
     6661 gtgagtcgta ttacgcgcgc agcggccgac catggcccaa cttgtttatt gcagcttata
     6721 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     6781 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg atctccggac
     6841 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     6901 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     6961 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggaagctgt aagcttgtcg
     7021 agaagtacta gaggatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     7081 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     7141 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     7201 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     7261 atcatgtctg gatctgatca ctgatatcgc ctaggagatc cgaaccagat aagtgaaatc
     7321 tagttccaaa ctattttgtc atttttaatt ttcgtattag cttacgacgc tacacccagt
     7381 tcccatctat tttgtcactc ttccctaaat aatccttaaa aactccattt ccacccctcc
     7441 cagttcccaa ctattttgtc cgcccacagc ggggcatttt tcttcctgtt atgtttttaa
     7501 tcaaacatcc tgccaactcc atgtgacaaa ccgtcatctt cggctacttt ttctctgtca
     7561 cagaatgaaa atttttctgt catctcttcg ttattaatgt ttgtaattga ctgaatatca
     7621 acgcttattt gcagcctgaa tggcgaatg
//
