LOCUS       sequence-246618-        5868 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses shRNA against human CNP under mouse U6 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5868)
  AUTHORS   Unlu I, Lu Y, Wang X
  TITLE     The cyclic phosphodiesterase CNP and RNA cyclase RtcA fine-tune 
            noncanonical XBP1 splicing during ER stress.
  JOURNAL   J Biol Chem. 2018 Dec 14;293(50):19365-19376. doi: 
            10.1074/jbc.RA118.004872. Epub 2018 Oct 24.
  PUBMED    30355738
REFERENCE   2  (bases 1 to 5868)
  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.RA118.004872"; journalName: "J Biol Chem"; date: 
            "2018-12-14- 14"; volume: "293"; issue: "50"; pages: "19365-19376"
COMMENT     Sequence Label: pB-puro-mU6-CNPshRNA2
FEATURES             Location/Qualifiers
     source          1..5868
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..314
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     233..253
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     repeat_region   716..778
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(844..863)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(845..863)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(870..887)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(870..886)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(879..901)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1028..1483
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1115..1134)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1325..1346
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1509..1613
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1614..1682,1683..2474)
                     /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(1832..1851)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2645..3233
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3134..3153
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3387..3404
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3521..3542
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3557..3562,3563..3580,3581..3587)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3595..3611
                     /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     3600..3622
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3619..3635
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3619..3635
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        3656..3674
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   3823..3857
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     polyA_signal    4244..4365
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4255..4274)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4309..4328
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(4699..5301)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MGTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4782..4802)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     regulatory      5298..5307
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(5321..5820)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(5369..5388)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     5762..5785
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
ORIGIN
        1 gatccgacgc cgccatctct aggcccgcgc cggccccctc gcacagactt gtgggagaag
       61 ctcggctact cccctgcccc ggttaatttg catataatat ttcctagtaa ctatagaggc
      121 ttaatgtgcg ataaaagaca gataatctgt tctttttaat actagctaca ttttacatga
      181 taggcttgga tttctataag agatacaaat actaaattat tattttaaaa aacagcacaa
      241 aaggaaactc accctaactg taaagtaatt gtgtgttttg agactataaa tatcccttgg
      301 agaaaagcct tgtttggagt acgctcaaca agatgtcgaa acatcttgtt gagcgtactc
      361 ctttttctcg agttaattaa cgagagcata atattgatat gtgccaaagt tgtttctgac
      421 tgactaataa gtataatttg tttctattat gtataggtta agctaattac ttattttata
      481 atacaacatg actgttttta aagtacaaaa taagtttatt tttgtaaaag agagaatgtt
      541 taaaagtttt gttactttat agaagaaatt ttgagttttt gttttttttt aataaataaa
      601 taaacataaa taaattgttt gttgaattta ttattagtat gtaagtgtaa atataataaa
      661 acttaatatc tattcaaatt aataaataaa cctcgatata cagaccgata aaacacatgc
      721 gtcaatttta cgcatgatta tctttaacgt acgtcacaat atgattatct ttctagggtt
      781 aaataatagt ttctaatttt tttattattc agcctgctgt cgtgaatacc gagctccaat
      841 tcgccctata gtgagtcgta ttacaattca ctggccgtcg ttttacaacg tcgtgactgg
      901 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
      961 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     1021 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1081 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1141 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1201 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1261 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1321 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1381 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     1441 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     1501 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     1561 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     1621 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     1681 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     1741 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     1801 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     1861 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     1921 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     1981 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     2041 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     2101 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     2161 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     2221 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     2281 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     2341 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     2401 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     2461 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     2521 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     2581 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     2641 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2701 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     2761 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     2821 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2881 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2941 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3001 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3061 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3121 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3181 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3241 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     3301 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     3361 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     3421 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     3481 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     3541 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     3601 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag ctcgaaatta
     3661 accctcacta aagggaacaa aagctggtac ctcgcgcgac ttggtttgcc attctttagc
     3721 gcgcgtcgcg tcacacagct tggccacaat gtggtttttg tcaaacgaag attctatgac
     3781 gtgtttaaag tttaggtcga gtaaagcgca aatctttttt aaccctagaa agatagtctg
     3841 cgtaaaattg acgcatgcat tcttgaaata ttgctctctc tttctaaata gcgcgaatcc
     3901 gtcgctgtgc atttaggaca tctcagtcgc cgcttggagc tcccgtgagg cgtgcttgtc
     3961 aatgcggtaa gtgtcactga ttttgaacta taacgaccgc gtgagtcaaa atgacgcatg
     4021 attatctttt acgtgacttt taagatttaa ctcatacgat aattatattg ttatttcatg
     4081 ttctacttac gtgataactt attatatata tattttcttg ttatagatat cgtgactaat
     4141 atataataaa atgggtagtt ctttagacga tgagcatatc ctctctgctc ttctgcaaag
     4201 cgatgacgag cttgttggct agcggcgcgc cgtcgagaca tgataagata cattgatgag
     4261 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     4321 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     4381 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
     4441 ctctacaaat gtggtatggc tgattatgat ctctagtcaa ggcactatac atcaaatatt
     4501 ccttattaac ccctttacaa attaaaaagc taaaggtaca caatttttga gcatagttat
     4561 taatagcaga cactctatgc ctgtgtggag taagaaaaaa cagtatgtta tgattataac
     4621 tgttatgcct acttataaag gttacagaat atttttccat aattttcttg tatagcagtg
     4681 cagctttttc tctagagctc aggcaccggg cttgcgggtc atgcaccagg tgcgcggtcc
     4741 ttcgggcacc tcgacgtcgg cggtgacggt gaagccgagc cgctcgtaga aggggaggtt
     4801 gcggggcgcg gaggtctcca ggaaggcggg caccccggcg cgctcggccg cctccactcc
     4861 ggggagcacg acggcgctgc ccagaccctt gccctggtgg tcgggcgaga cgccgacggt
     4921 ggccaggaac cacgcgggct ccttgggccg gtgcggcgcc aggaggcctt ccatctgttg
     4981 ctgcgcggcc agccgggaac cgctcaactc ggccatgcgc gggccgatct cggcgaacac
     5041 cgcccccgct tcgacgctct ccggcgtggt ccagaccgcc accgcggcgc cgtcgtccgc
     5101 gacccacacc ttgccgatgt cgagcccgac gcgcgtgagg aagagttctt gcagctcggt
     5161 gacccgctcg atgtggcggt ccgggtcgac ggtgtggcgc gtggcggggt agtcggcgaa
     5221 cgcggcggcg agggtgcgta cggcccgggg gacgtcgtcg cgggtggcga ggcgcaccgt
     5281 gggcttgtac tcggtcccca tggtggcgtt ggctgcaggt cgaaaggccc ggagatgagg
     5341 aagaggagaa cagcgcggca gacgtgcgct tttgaagcgt gcagaatgcc gggcctccgg
     5401 aggaccttcg ggcgcccgcc ccgcccctga gcccgcccct gagcccgccc ccggacccac
     5461 cccttcccag cctctgagcc cagaaagcga aggagcaaag ctgctattgg ccgctgcccc
     5521 aaaggcctac ccgcttccat tgctcagcgg tgctgtccat ctgcacgaga ctagtgagac
     5581 gtgctacttc catttgtcac gtcctgcacg acgcgagctg cggggcgggg gggaacttcc
     5641 tgactagggg aggagtagaa ggtggcgcga aggggccacc aaagaacgga gccggttggc
     5701 gcctaccggt ggatgtggaa tgtgtgcgag gccagaggcc acttgtgtag cgccaagtgc
     5761 ccagcggggc tgctaaagcg catgctccag actgccttgg gaaaagcgcc tcccctaccc
     5821 ggtagaatta attcgatggc cgcggtccac actcgcccca acgcccgg
//
