LOCUS       pAAV_GFAP_CaBLAM        6605 bp    DNA     circular SYN 07-MAR-2025
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 6605)
  AUTHORS   Hochgeschwender
  TITLE     Direct Submission
  JOURNAL   Exported Jul 25, 2025 from SnapGene 8.1.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6605
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     rep_origin      153..741
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     642..661
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
                     /note="color: black; sequence: GGGAAACGCCTGGTATCTTT"
     repeat_region   803..932
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
                     /note="color: #ffcc99"
     promoter        952..1632
                     /label=GFAP promoter
                     /note="color: #999999"
     primer_bind     complement(1297..1308)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
                     /note="color: black; sequence: ATAATACCGCGCCACATAGC"
     primer_bind     1707..1723
                     /label=C883
                     /note="color: black; sequence: 
                     gagactctGGATccATGgtgagcaagggc; added: 2024-03-04"
     CDS             1709..2386
                     /codon_start=1
                     /label=mNG_d10c
                     /note="color: #00ff00"
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFT"
     regulatory      1964..1973
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     primer_bind     2171..2190
                     /label=C644
                     /note="color: black; sequence: taccccaacgacaaaaccat; added:
                     2022-10-18"
     primer_bind     complement(2171..2190)
                     /label=C646
                     /note="color: black; sequence: atggttttgtcgttggggta; added:
                     2022-10-18"
     CDS             2387..2392
                     /codon_start=1
                     /label=GF_linker
                     /note="color: #808080"
                     /translation="GF"
     CDS             2393..2851
                     /codon_start=1
                     /label=ekl9h aa 6-156
                     /note="color: #00ffff"
                     /translation="EDFVGDWEQTAAYNLDQVLEQGGVSSVLQTLAVSVTPIQRIVRSG
                     ENGLKIDIHVIIPYEGLSADQMAHIEEVFKVVYPVDDHHFKVIMEYGTLVIDGVTPNML
                     NYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLITPEGSMLFRVTIN"
     primer_bind     2399..2417
                     /label=C608
                     /note="color: black; sequence: TTTCGTTGGGGACTGGGAAC; added:
                     2024-03-04"
     primer_bind     complement(2456..2479)
                     /label=C884
                     /note="color: black; sequence: 
                     agagtctcCTGCAGAACAGAAGAAACACCACC; added: 2024-03-04"
     primer_bind     2473..2490
                     /label=C885
                     /note="color: black; sequence: gagactctCTGCAGACCCTGGCTGT; 
                     added: 2024-03-04"
     primer_bind     2830..2849
                     /label=C650
                     /note="color: black; sequence: GCTGTTCCGTGTTACCATCA; added:
                     2022-10-18"
     CDS             2852..2863
                     /codon_start=1
                     /label=SASS linker
                     /note="color: #808080"
                     /translation="SASS"
     CDS             2864..2929
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /note="color: #ff6600"
                     /translation="DSSRRKWNKTGHAVRAIGRLSS"
     CDS             2930..2974
                     /codon_start=1
                     /label=5xGGSLinker
                     /note="color: #808080"
                     /translation="GGSGGSGGSGGSGGS"
     CDS             2975..3424
                     /codon_start=1
                     /label=Calmodulin
                     /note="color: #ff6600"
                     /translation="NLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMRSLGQNP
                     TEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGIGYISA
                     AELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     CDS             3425..3433
                     /codon_start=1
                     /label=SPA linker
                     /note="color: #808080"
                     /translation="SPA"
     CDS             3434..3469
                     /codon_start=1
                     /product="sequence from eBlocks_11-15-2020 well D12 
                     APD3A6_LLEEISN"
                     /label=LLEEISN peptide
                     /note="color: #00ffff"
                     /translation="VTGYRLLEEISN"
     primer_bind     complement(3445..3469)
                     /label=C886
                     /note="color: black; sequence: 
                     agagtctcctcgagttattctaaGTTAGAGATTTCTTCGAGCAGACGG; added: 
                     2024-03-04"
     misc_feature    3534..4122
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
                     /note="color: #ffffff"
     primer_bind     complement(3587..3607)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
                     /note="color: black; sequence: CATAGCGTAAAAGGAGCAACA"
     CDS             complement(4005..4016)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /note="color: #cc99b2
                     Cleavage site after base 4004"
                     /translation="IEGR"
     polyA_signal    4154..4630
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(4353..4372)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
                     /note="color: black; sequence: CCAGCTTGGTTCCCAATAGA"
     repeat_region   4670..4810
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4"
     repeat_region   4670..4799
                     /label=AAV2 ITR
                     /note="color: #ffcc99"
     rep_origin      4885..5340
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     complement(4972..4991)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
                     /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG"
     primer_bind     5182..5203
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
                     /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG"
     primer_bind     complement(5357..5376)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
                     /note="color: black; sequence: CGGCATCAGAGCAGATTGTA"
     primer_bind     5476..5498
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
                     /note="color: black; sequence: CCGGGAGCTGCATGTGTCAGAGG"
     primer_bind     complement(5536..5554)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
                     /note="color: black; sequence: AATAGGCGTATCACGAGGC"
     promoter        5622..5726
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             join(5727..5795,5796..6587)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 5727 .. 5795 / #ccffcc / signal sequence
                        2: 5796 .. 6587 / #ccffcc
                     Cleavage site after base 5795"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5945..5964)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
                     /note="color: black; sequence: ATAATACCGCGCCACATAGC"
ORIGIN
        1 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
       61 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
      121 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
      181 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
      241 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
      301 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
      361 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
      421 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
      481 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
      541 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
      601 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
      661 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
      721 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
      781 tgctggcctt ttgctcacat gtcctgcagg cagctgcgcg ctcgctcgct cactgaggcc
      841 gcccgggcgt cgggcgacct ttggtcgccc ggcctcagtg agcgagcgag cgcgcagaga
      901 gggagtggcc aactccatca ctaggggttc ctgcggccgc acgcgtgatc taacatatcc
      961 tggtgtggag taggggacgc tgctctgaca gaggctcggg ggcctgagct ggctctgtga
     1021 gctggggagg aggcagacag ccaggccttg tctgcaagca gacctggcag cattgggctg
     1081 gccgcccccc agggcctcct cttcatgccc agtgaatgac tcaccttggc acagacacaa
     1141 tgttcggggt gggcacagtg cctgcttccc gccgcacccc agcccccctc aaatgccttc
     1201 cgagaagccc attgagcagg gggcttgcat tgcaccccag cctgacagcc tggcatcttg
     1261 ggataaaagc agcacagccc cctaggggct gcccttgctg tgtggcgcca ccggcggtgg
     1321 agaacaaggc tctattcagc ctgtgcccag gaaaggggat caggggatgc ccaggcatgg
     1381 acagtgggtg gcaggggggg agaggagggc tgtctgcttc ccagaagtcc aaggacacaa
     1441 atgggtgagg ggagagctct ccccatagct gggctgcggc ccaaccccac cccctcaggc
     1501 tatgccaggg ggtgttgcca ggggcacccg ggcatcgcca gtctagccca ctccttcata
     1561 aagccctcgc atcccaggag cgagcagagc cagagcaggt tggagaggag acgcatcacc
     1621 tccgctgctc gcggggtcta gagtcgacaa gcttggatct cgacaagctg ggcgcgccaa
     1681 tattatgcta gcgctaccgg tcgccaccat ggtgagcaag ggcgaggagg ataacatggc
     1741 ctctctccca gcgacacatg agttacacat ctttggctcc atcaacggtg tggactttga
     1801 catggtgggt cagggcaccg gcaatccaaa tgatggttat gaggagttaa acctgaagtc
     1861 caccaagggt gacctccagt tctccccctg gattctggtc cctcatatcg ggtatggctt
     1921 ccatcagtac ctgccctacc ctgacgggat gtcgcctttc caggccgcca tggtagatgg
     1981 ctccggatac caagtccatc gcacaatgca gtttgaagat ggtgcctccc ttactgttaa
     2041 ctaccgctac acctacgagg gaagccacat caaaggagag gcccaggtga aggggactgg
     2101 tttccctgct gacggtcctg tgatgaccaa ctcgctgacc gctgcggact ggtgcaggtc
     2161 gaagaagact taccccaacg acaaaaccat catcagtacc tttaagtgga gttacaccac
     2221 tggaaatggc aagcgctacc ggagcactgc gcggaccacc tacacctttg ccaagccaat
     2281 ggcggctaac tatctgaaga accagccgat gtacgtgttc cgtaagacgg agctcaagca
     2341 ctccaagacc gagctcaact tcaaggagtg gcaaaaggcc tttaccggat ttgaagactt
     2401 cgttggtgac tgggaacaga ccgctgctta caacctggac caggttctgg aacagggtgg
     2461 tgtttcttct gttctgcaga ccctggctgt ttctgttacc cctatccagc gtatcgttcg
     2521 ttctggtgaa aacggtctga agatcgacat ccatgttatc atcccttacg aaggtctgtc
     2581 tgctgaccag atggctcata tcgaagaagt tttcaaggtt gtttaccctg ttgacgacca
     2641 tcatttcaag gttatcatgg aatacggtac cctggttatc gacggtgtta cccctaacat
     2701 gctgaactac ttcggtcgtc cttacgaagg tatcgctgtt ttcgacggta agaagatcac
     2761 cgttaccggt accctgtgga acggtaacaa gatcatcgac gaacgtctga tcacccctga
     2821 gggttctatg ctgttccgtg ttaccatcaa ctctgcttct tctgactctt ctaggcgtaa
     2881 gtggaacaag accggtcatg ctgttcgtgc tatcggtcgt ctgtcttctg gtggatccgg
     2941 tggtagtggt ggatccggag gttctggtgg atccaacctg cctgaccagc tgaccgaaga
     3001 acagatcgct gaattcaagg aagagttctc tctgttcgat aaagatggag acggtaccat
     3061 caccaccaag gaactgggta ccgtgatgcg ttctctgggt cagaacccta ccgaagctga
     3121 actgcaagac atgatcaacg aagttgacgc tgacggtgac ggtaccatcg acttccctga
     3181 attcctgacc atgatggctc gtaagatgaa gtacagggac accgaagagg aaatccgtga
     3241 agctttcggc gttttcgaca aggacggtat cggttacatc tctgctgctg aactgcgtca
     3301 tgttatgacc aacctgggtg aaaagctgac cgacgaagaa gttgacgaaa tgatccgtga
     3361 agctgacatc gacggtgacg gtcaggttaa ctacgaagaa ttcgttcaga tgatgaccgc
     3421 taagtctcct gctgttaccg gttaccgtct gctcgaagaa atctctaacc tcgagtaaca
     3481 tatgcacgcg tcaattgctc gaggctagcg aattcgatat caagcttatc gataatcaac
     3541 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     3601 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     3661 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     3721 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     3781 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     3841 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     3901 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctatg
     3961 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     4021 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     4081 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgagcgctgc
     4141 tcgagagatc tacgggtggc atccctgtga cccctcccca gtgcctctcc tggccctgga
     4201 agttgccact ccagtgccca ccagccttgt cctaataaaa ttaagttgca tcattttgtc
     4261 tgactaggtg tccttctata atattatggg gtggaggggg gtggtatgga gcaaggggca
     4321 agttgggaag acaacctgta gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt
     4381 ggcacaatct tggctcactg caatctccgc ctcctgggtt caagcgattc tcctgcctca
     4441 gcctcccgag ttgttgggat tccaggcatg catgaccagg ctcagctaat ttttgttttt
     4501 ttggtagaga cggggtttca ccatattggc caggctggtc tccaactcct aatctcaggt
     4561 gatctaccca ccttggcctc ccaaattgct gggattacag gcgtgaacca ctgctccctt
     4621 ccctgtcctt ctgattttgt aggtaaccac gtgcggaccg agcggccgca ggaaccccta
     4681 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
     4741 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc
     4801 tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     4861 cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     4921 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg
     4981 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     5041 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     5101 tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     5161 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta
     5221 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa
     5281 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     5341 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     5401 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     5461 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     5521 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     5581 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     5641 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     5701 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     5761 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     5821 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     5881 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     5941 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     6001 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     6061 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     6121 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     6181 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     6241 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     6301 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     6361 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     6421 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     6481 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     6541 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     6601 tttac
//
