LOCUS       sequence-81960-e        4654 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shRNA silencing plasmid with eGFP transfection marker .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4654)
  AUTHORS   Schnizler MK, Schnizler K, Zha XM, Hall DD, Wemmie JA, Hell JW, 
            Welsh MJ
  TITLE     The cytoskeletal protein alpha-actinin regulates acid-sensing ion 
            channel 1a through a C-terminal interaction.
  JOURNAL   J Biol Chem. 2009 Jan 30;284(5):2697-705. doi: 
            10.1074/jbc.M805110200. Epub 2008 Nov 21.
  PUBMED    19028690
REFERENCE   2  (bases 1 to 4654)
  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: 
            "10.1074/jbc.M805110200"; journalName: "J Biol Chem"; date: 
            "2009-01-30- 30"; volume: "284"; issue: "5"; pages: "2697-705"
COMMENT     Sequence Label: pSIL-eGFP
FEATURES             Location/Qualifiers
     source          1..4654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        84..387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        388..591
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     542..562
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      630..639
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(636..638,639..641,642..1358)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYKY"
     primer_bind     complement(681..702)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(942..961)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1289..1310
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1367..1383
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1380..1693
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     1612..1632
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     1705..1721
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1706..1722
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1756..1772)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1809..1829)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1809..1827)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1848..1864)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1848..1864)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1861..1883)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1872..1888
                     /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)."
     promoter        complement(join(1896..1902,1903..1920,1921..1926))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1941..1962
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2079..2096)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2250..2838)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2330..2349)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3009..3800,3801..3869))
                     /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     3632..3651
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3870..3974)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4000..4455)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4137..4158)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4349..4368
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4582..4604
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4596..4613
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4597..4613
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4623..4642
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4623..4641
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ctaaccgtat taccgccatg cattagttat taatagtaat caattacggg gtcattagtt
       61 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      121 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      181 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
      241 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      301 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      361 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
      421 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
      481 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
      541 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg
      601 aaccgtcaga tccgctagcg ctaccggtcg ccaccatggt gagcaagggc gaggagctgt
      661 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
      721 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
      781 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
      841 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
      901 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
      961 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     1021 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     1081 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     1141 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     1201 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga
     1261 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     1321 ggatcactct cggcatggac gagctgtaca agtactaggt acccgctcta gaactagtgg
     1381 atccgacgcc gccatctcta ggcccgcgcc ggccccctcg cacagacttg tgggagaagc
     1441 tcggctactc ccctgccccg gttaatttgc atataatatt tcctagtaac tatagaggct
     1501 taatgtgcga taaaagacag ataatctgtt ctttttaata ctagctacat tttacatgat
     1561 aggcttggat ttctataaga gatacaaata ctaaattatt attttaaaaa acagcacaaa
     1621 aggaaactca ccctaactgt aaagtaattg tgtgttttga gactataaat atcccttgga
     1681 gaaaagcctt gtttgggccc cccctcgagg tcgacggtat cgataagctt gatatcgaat
     1741 tcctgcagcc cgggggatcc actagttcta gagcggccgc caccgcggtg gagctccagc
     1801 ttttgttccc tttagtgagg gttaattgcg cgcttggcgt aatcatggtc atagctgttt
     1861 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     1921 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     1981 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     2041 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     2101 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     2161 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     2221 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     2281 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     2341 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     2401 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     2461 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     2521 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     2581 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     2641 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt
     2701 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     2761 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     2821 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     2881 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     2941 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     3001 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     3061 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     3121 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     3181 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     3241 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     3301 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     3361 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     3421 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     3481 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     3541 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     3601 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     3661 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     3721 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     3781 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     3841 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     3901 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     3961 ataggggttc cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt
     4021 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     4081 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     4141 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     4201 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     4261 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     4321 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     4381 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     4441 cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg
     4501 tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa
     4561 gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgagcgcg
     4621 cgtaatacga ctcactatag ggcgaattgg gtac
//
