LOCUS       sequence-149133-        6562 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bicistronic construct expressing PACSIN1 shRNA and HA-PACSIN1-rescue
            construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6562)
  AUTHORS   Anggono V, Koc-Schmitz Y, Widagdo J, Kormann J, Quan A, Chen CM, 
            Robinson PJ, Choi SY, Linden DJ, Plomann M, Huganir RL
  TITLE     PICK1 interacts with PACSIN to regulate AMPA receptor 
            internalization and cerebellar long-term depression.
  JOURNAL   Proc Natl Acad Sci U S A. 2013 Aug 20;110(34):13976-81. doi: 
            10.1073/pnas.1312467110. Epub 2013 Aug 5.
  PUBMED    23918399
REFERENCE   2  (bases 1 to 6562)
  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"; doi: 
            "10.1073/pnas.1312467110"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2013-08-20- 20"; volume: "110"; issue: "34"; pages: 
            "13976-81"
COMMENT     Sequence Label: pRK5-H1-shRNA-CMV-HA-PACSIN1
FEATURES             Location/Qualifiers
     source          1..6562
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      963..972
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             975..1001
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2391..2400
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2404..2538
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2441..2460)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2495..2514
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2611..2968
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2611..2802
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2634..2654)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2819..2954
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2988..3005)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2988..3004)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2997..3019)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3217..3672
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3304..3323)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3514..3535
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3689..3708)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3843..3865
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3918..3936)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4004..4108
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4109..4177,4178..4969)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRDDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4327..4346)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5140..5728
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5629..5648
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5790..5809
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5790..5808
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     5841..5857
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5841..5857
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        5877..6091
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     primer_bind     6012..6031
                     /label=H1
                     /note="Human H1 promoter, forward primer"
     primer_bind     6298..6315
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6432..6453
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6468..6473,6474..6491,6492..6498)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6506..6522
                     /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)."
     primer_bind     6511..6533
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6530..6546
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6530..6546
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataac cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attcccgaac cgacagtcgg tctcttcacc aaggccattc
      961 gcgccaccat gggctacccc tatgatgtgc ctgactacgc agatctcaat ggtggtggtg
     1021 gtgggtcgac catgtctggc tcctacgatg aggcctcaga ggagatcaca gatagcttct
     1081 gggaggtggg gaactacaag cggacggtga agcgcatcga cgatgggcac cgcctgtgca
     1141 acgacctcat gagctgcgtg caggagcgcg ccaagatcga gaaggcatac gcgcagcagc
     1201 tcaccgactg ggccaagcgc tggcgccaac ttattgaaaa gggtcctcag tatggcagcc
     1261 tggagcgggc gtggggcgcc atgatgacag aagcagataa ggtcagcgag ctgcaccagg
     1321 aggtgaagaa cagcctgctg aatgaggacc tggagaaagt caagaactgg cagaaggatg
     1381 cctatcacaa gcagatcatg ggtggcttca aggagacgaa agaggccgag gatggcttcc
     1441 gaaaggccca gaagccctgg gctaaaaaga tgaaggagct agaggcggcc aagaaggcct
     1501 atcacttggc ttgtaaggag gaaaggctgg ccatgacccg ggagatgaac agtaagacag
     1561 agcagtcggt cacccctgaa cagcagaaga aacttgtgga caaagtggac aaatgcagac
     1621 aggatgtgca aaagactcag gagaagtatg agaaggtgct ggaagatgtg ggcaagacca
     1681 caccacagta catggagggc atggagcagg tgtttgagca gtgccagcag tttgaggaga
     1741 agcggctggt cttcctgaag gaagtcctgc tggatatcaa acggcatctc aacctagcgg
     1801 agaacagcag ctacatgcat gtctaccgag aactggagca ggccatccgg ggggccgatg
     1861 cccaggagga cctcaggtgg ttccgcagca ccagtggccc cgggatgccc atgaactggc
     1921 cgcagttcga ggagtggaac ccagacctcc cgcacaccac tgccaagaag gagaaacagc
     1981 ctaagaaggc agagggggcc accctgagca atgccactgg ggctgtagaa tccacatccc
     2041 aggctgggga ccgtggcagt gttagcagct atgaccgagg ccaaacatat gccaccgagt
     2101 ggtcagacga tgagagcgga aaccccttcg ggggcaatga ggccaatggt ggcgccaacc
     2161 ccttcgagga tgatgccaag ggagttcgtg tacgggcact ctatgactac gacggtcagg
     2221 agcaggatga gctcagcttc aaggccggag atgagctcac caagctcgga gaggaagacg
     2281 aacagggttg gtgccgcggg cggctggaca gcggacagct gggcctctat cctgccaact
     2341 acgttgaggc tatataggcg gccgctaagt aagtaaggat ccccagcttg gccgccatgg
     2401 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     2461 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     2521 tcttatcatg tctggatcga tcgggaatta attcggcgca gcaccatggc ctgaaataac
     2581 ctctgaaaga ggaacttggt taggtacctt ctgaggcgga aagaaccagc tgtggaatgt
     2641 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     2701 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     2761 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     2821 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     2881 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     2941 cttttttgga ggcctaggct tttgcaaaaa gctgttaaca gcttggcact ggccgtcgtt
     3001 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     3061 ccccccttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     3121 ttgcgtagcc tgaatggcga atggcgcctg atgcggtatt ttctccttac gcatctgtgc
     3181 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     3241 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     3301 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     3361 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     3421 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     3481 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     3541 cactcaaccc tatctcgggc tattcttttg atttataagg gattttgccg atttcggcct
     3601 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     3661 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     3721 aactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc
     3781 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     3841 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagta
     3901 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     3961 aatggtttct tagacgtcag gtttcacttt tcggggaaat gtgcgcggaa cccctatttg
     4021 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     4081 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     4141 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4201 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     4261 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     4321 agttctgcta tgtggcgcgg tattatcccg tgatgacgcc gggcaagagc aactcggtcg
     4381 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     4441 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     4501 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     4561 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     4621 accaaacgac gagcgtgaca ccacgatgcc agcagcaatg gcaacaacgt tgcgcaaact
     4681 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     4741 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     4801 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     4861 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     4921 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     4981 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     5041 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     5101 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5161 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     5221 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     5281 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     5341 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     5401 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5461 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     5521 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     5581 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     5641 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     5701 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     5761 ggccttttgc tggccttttg ctcacatgtt aatacgactc actatagggc gaattggagc
     5821 tccaccgcgg tggcggccgc tctagaacta gtggatcccc cgggctgcag gaattcgaac
     5881 gctgacgtca tcaacccgct ccaaggaatc gcgggcccag tgtcactagg cgggaacacc
     5941 cagcgcgcgt gcgccctggc aggaagatgg ctgtgaggga caggggagtg gcgccctgca
     6001 atatttgcat gtcgctatgt gttctgggaa atcaccataa acgtgaaatg tctttggatt
     6061 tgggaatctt ataagttctg tatgagacca cagatccccg cgccagctca tcgagaaatt
     6121 aagagatttc tcgatgagct ggcgcttttt tctcgagggg gggcccggta ccagcttttg
     6181 ttccctttat gagggttaaa catgttcttt cctgcgttat cccctgattc tgtggataac
     6241 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     6301 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     6361 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     6421 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
     6481 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
     6541 tatgaccatg attacgaatt aa
//
