LOCUS       sequence-246621-        5871 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses shRNA against human RtcA under mouse U6 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5871)
  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 5871)
  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-RtcAshRNA1
FEATURES             Location/Qualifiers
     source          1..5871
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      66..75
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(89..588)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(137..156)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     530..553
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     promoter        637..950
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     869..889
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     repeat_region   1355..1417
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(1483..1502)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1484..1502)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1509..1526)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1509..1525)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1518..1540)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1667..2122
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1754..1773)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1964..1985
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2148..2252
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2253..2321,2322..3113)
                     /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(2471..2490)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3284..3872
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3773..3792
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4026..4043
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4160..4181
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4196..4201,4202..4219,4220..4226)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4234..4250
                     /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     4239..4261
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4258..4274
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4258..4274
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        4295..4313
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   4462..4496
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     polyA_signal    4883..5004
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4894..4913)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4948..4967
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(5338..5871,1..69))
                     /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(5421..5441)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
ORIGIN
        1 ggtgcgtacg gcccggggga cgtcgtcgcg ggtggcgagg cgcaccgtgg gcttgtactc
       61 ggtccccatg gtggcgttgg ctgcaggtcg aaaggcccgg agatgaggaa gaggagaaca
      121 gcgcggcaga cgtgcgcttt tgaagcgtgc agaatgccgg gcctccggag gaccttcggg
      181 cgcccgcccc gcccctgagc ccgcccctga gcccgccccc ggacccaccc cttcccagcc
      241 tctgagccca gaaagcgaag gagcaaagct gctattggcc gctgccccaa aggcctaccc
      301 gcttccattg ctcagcggtg ctgtccatct gcacgagact agtgagacgt gctacttcca
      361 tttgtcacgt cctgcacgac gcgagctgcg gggcgggggg gaacttcctg actaggggag
      421 gagtagaagg tggcgcgaag gggccaccaa agaacggagc cggttggcgc ctaccggtgg
      481 atgtggaatg tgtgcgaggc cagaggccac ttgtgtagcg ccaagtgccc agcggggctg
      541 ctaaagcgca tgctccagac tgccttggga aaagcgcctc ccctacccgg tagaattaat
      601 tcgatggccg cggtccacac tcgccccaac gcccgggatc cgacgccgcc atctctaggc
      661 ccgcgccggc cccctcgcac agacttgtgg gagaagctcg gctactcccc tgccccggtt
      721 aatttgcata taatatttcc tagtaactat agaggcttaa tgtgcgataa aagacagata
      781 atctgttctt tttaatacta gctacatttt acatgatagg cttggatttc tataagagat
      841 acaaatacta aattattatt ttaaaaaaca gcacaaaagg aaactcaccc taactgtaaa
      901 gtaattgtgt gttttgagac tataaatatc ccttggagaa aagccttgtt tgtggcacca
      961 cagatcgatt atacaagagt ataatcgatc tgtggtgcca tttttctcga gttaattaac
     1021 gagagcataa tattgatatg tgccaaagtt gtttctgact gactaataag tataatttgt
     1081 ttctattatg tataggttaa gctaattact tattttataa tacaacatga ctgtttttaa
     1141 agtacaaaat aagtttattt ttgtaaaaga gagaatgttt aaaagttttg ttactttata
     1201 gaagaaattt tgagtttttg ttttttttta ataaataaat aaacataaat aaattgtttg
     1261 ttgaatttat tattagtatg taagtgtaaa tataataaaa cttaatatct attcaaatta
     1321 ataaataaac ctcgatatac agaccgataa aacacatgcg tcaattttac gcatgattat
     1381 ctttaacgta cgtcacaata tgattatctt tctagggtta aataatagtt tctaattttt
     1441 ttattattca gcctgctgtc gtgaataccg agctccaatt cgccctatag tgagtcgtat
     1501 tacaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     1561 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     1621 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc
     1681 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     1741 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     1801 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     1861 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     1921 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     1981 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2041 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     2101 aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     2161 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     2221 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     2281 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     2341 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     2401 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     2461 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     2521 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     2581 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     2641 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     2701 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     2761 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     2821 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     2881 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     2941 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     3001 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     3061 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     3121 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     3181 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     3241 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     3301 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     3361 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     3421 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     3481 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     3541 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     3601 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     3661 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     3721 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     3781 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     3841 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     3901 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     3961 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     4021 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc
     4081 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     4141 gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac
     4201 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag
     4261 gaaacagcta tgaccatgat tacgccaagc tcgaaattaa ccctcactaa agggaacaaa
     4321 agctggtacc tcgcgcgact tggtttgcca ttctttagcg cgcgtcgcgt cacacagctt
     4381 ggccacaatg tggtttttgt caaacgaaga ttctatgacg tgtttaaagt ttaggtcgag
     4441 taaagcgcaa atctttttta accctagaaa gatagtctgc gtaaaattga cgcatgcatt
     4501 cttgaaatat tgctctctct ttctaaatag cgcgaatccg tcgctgtgca tttaggacat
     4561 ctcagtcgcc gcttggagct cccgtgaggc gtgcttgtca atgcggtaag tgtcactgat
     4621 tttgaactat aacgaccgcg tgagtcaaaa tgacgcatga ttatctttta cgtgactttt
     4681 aagatttaac tcatacgata attatattgt tatttcatgt tctacttacg tgataactta
     4741 ttatatatat attttcttgt tatagatatc gtgactaata tataataaaa tgggtagttc
     4801 tttagacgat gagcatatcc tctctgctct tctgcaaagc gatgacgagc ttgttggcta
     4861 gcggcgcgcc gtcgagacat gataagatac attgatgagt ttggacaaac cacaactaga
     4921 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     4981 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
     5041 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtatggct
     5101 gattatgatc tctagtcaag gcactataca tcaaatattc cttattaacc cctttacaaa
     5161 ttaaaaagct aaaggtacac aatttttgag catagttatt aatagcagac actctatgcc
     5221 tgtgtggagt aagaaaaaac agtatgttat gattataact gttatgccta cttataaagg
     5281 ttacagaata tttttccata attttcttgt atagcagtgc agctttttct ctagagctca
     5341 ggcaccgggc ttgcgggtca tgcaccaggt gcgcggtcct tcgggcacct cgacgtcggc
     5401 ggtgacggtg aagccgagcc gctcgtagaa ggggaggttg cggggcgcgg aggtctccag
     5461 gaaggcgggc accccggcgc gctcggccgc ctccactccg gggagcacga cggcgctgcc
     5521 cagacccttg ccctggtggt cgggcgagac gccgacggtg gccaggaacc acgcgggctc
     5581 cttgggccgg tgcggcgcca ggaggccttc catctgttgc tgcgcggcca gccgggaacc
     5641 gctcaactcg gccatgcgcg ggccgatctc ggcgaacacc gcccccgctt cgacgctctc
     5701 cggcgtggtc cagaccgcca ccgcggcgcc gtcgtccgcg acccacacct tgccgatgtc
     5761 gagcccgacg cgcgtgagga agagttcttg cagctcggtg acccgctcga tgtggcggtc
     5821 cgggtcgacg gtgtggcgcg tggcggggta gtcggcgaac gcggcggcga g
//
