LOCUS       sequence-318932-        7080 bp    DNA     circular SYN 15-APR-2025
DEFINITION  SNX9 shRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7080)
  AUTHORS   Roman-Fernandez A, Roignot J, Sandilands E, Nacke M, Mansour MA, 
            McGarry L, Shanks E, Mostov KE, Bryant DM
  TITLE     The phospholipid PI(3,4)P2 is an apical identity determinant.
  JOURNAL   Nat Commun. 2018 Nov 28;9(1):5041. doi: 10.1038/s41467-018-07464-8.
  PUBMED    30487552
REFERENCE   2  (bases 1 to 7080)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-11-28"; volume: "9(1):5041. doi"; pages: " 
            10.1038/s41467-018-07464-8"
COMMENT     Sequence Label: pLKO.1 Puro shRNA SNX9 KD1
FEATURES             Location/Qualifiers
     source          1..7080
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    158..275
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        324..834
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     753..771
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             856..1455
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(856..875)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1352..1372
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1583..1816
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1888..2009
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1925..1944)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1979..1998
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2049..2184
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2204..2223)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2205..2223)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2233..2250)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2233..2249)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2242..2264)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2391..2846
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2478..2497)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2688..2709
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2872..2976
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2977..3045,3046..3837)
                     /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(3195..3214)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4008..4596
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4497..4516
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4750..4767
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4884..4905
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4920..4925,4926..4943,4944..4950)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4958..4974
                     /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     4963..4985
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4982..4998
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4982..4998
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5017..5037
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5019..5037
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5065..5291
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5292..5472
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5519..5644
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6137..6370
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6555..6599
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        join(6897..7080,1..65)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6897..6917
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     join(7068..7080,1..7)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
ORIGIN
        1 ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg aaaggacgaa
       61 acaccggtcc cttctcttcc agataactcg agttatctgg aagagaaggg atttttgaat
      121 tctcgacctc gagacaaatg gcagtattca tccacaattt taaaagaaaa ggggggattg
      181 gggggtacag tgcaggggaa agaatagtag acataatagc aacagacata caaactaaag
      241 aattacaaaa acaaattaca aaaattcaaa attttcgggt ttattacagg gacagcagag
      301 atccactttg gccgcggctc gagggggttg gggttgcgcc ttttccaagg cagccctggg
      361 tttgcgcagg gacgcggctg ctctgggcgt ggttccggga aacgcagcgg cgccgaccct
      421 gggtctcgca cattcttcac gtccgttcgc agcgtcaccc ggatcttcgc cgctaccctt
      481 gtgggccccc cggcgacgct tcctgctccg cccctaagtc gggaaggttc cttgcggttc
      541 gcggcgtgcc ggacgtgaca aacggaagcc gcacgtctca ctagtaccct cgcagacgga
      601 cagcgccagg gagcaatggc agcgcgccga ccgcgatggg ctgtggccaa tagcggctgc
      661 tcagcagggc gcgccgagag cagcggccgg gaaggggcgg tgcgggaggc ggggtgtggg
      721 gcggtagtgt gggccctgtt cctgcccgcg cggtgttccg cattctgcaa gcctccggag
      781 cgcacgtcgg cagtcggctc cctcgttgac cgaatcaccg acctctctcc ccagggggat
      841 ccaccggagc ttaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
      901 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
      961 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     1021 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     1081 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     1141 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     1201 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     1261 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     1321 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
     1381 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     1441 aagcccggtg cctgacgccc gccccacgac ccgcagcgcc cgaccgaaag gagcgcacga
     1501 ccccatgcat cggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     1561 tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatctg
     1621 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     1681 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     1741 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     1801 tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca
     1861 aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa tggttacaaa
     1921 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     1981 ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc ccgcccctaa
     2041 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     2101 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     2161 agtgaggagg cttttttgga ggcctaggga cgtacccaat tcgccctata gtgagtcgta
     2221 ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     2281 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     2341 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg acgcgccctg
     2401 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     2461 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     2521 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     2581 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     2641 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     2701 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     2761 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     2821 taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc
     2881 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     2941 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     3001 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     3061 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     3121 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     3181 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     3241 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     3301 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     3361 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     3421 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     3481 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     3541 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     3601 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     3661 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     3721 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     3781 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     3841 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     3901 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     3961 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     4021 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     4081 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     4141 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     4201 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     4261 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     4321 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     4381 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     4441 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     4501 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     4561 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     4621 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     4681 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     4741 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct
     4801 ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa
     4861 gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct
     4921 ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac
     4981 acaggaaaca gctatgacca tgattacgcc aagcgcgcaa ttaaccctca ctaaagggaa
     5041 caaaagctgg agctgcaagc ttaatgtagt cttatgcaat actcttgtag tcttgcaaca
     5101 tggtaacgat gagttagcaa catgccttac aaggagagaa aaagcaccgt gcatgccgat
     5161 tggtggaagt aaggtggtac gatcgtgcct tattaggaag gcaacagacg ggtctgacat
     5221 ggattggacg aaccactgaa ttgccgcatt gcagagatat tgtatttaag tgcctagctc
     5281 gatacataaa cgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     5341 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     5401 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     5461 aaatctctag cagtggcgcc cgaacaggga cttgaaagcg aaagggaaac cagaggagct
     5521 ctctcgacgc aggactcggc ttgctgaagc gcgcacggca agaggcgagg ggcggcgact
     5581 ggtgagtacg ccaaaaattt tgactagcgg aggctagaag gagagagatg ggtgcgagag
     5641 cgtcagtatt aagcggggga gaattagatc gcgatgggaa aaaattcggt taaggccagg
     5701 gggaaagaaa aaatataaat taaaacatat agtatgggca agcagggagc tagaacgatt
     5761 cgcagttaat cctggcctgt tagaaacatc agaaggctgt agacaaatac tgggacagct
     5821 acaaccatcc cttcagacag gatcagaaga acttagatca ttatataata cagtagcaac
     5881 cctctattgt gtgcatcaaa ggatagagat aaaagacacc aaggaagctt tagacaagat
     5941 agaggaagag caaaacaaaa gtaagaccac cgcacagcaa gcggccgctg atcttcagac
     6001 ctggaggagg agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa
     6061 aaattgaacc attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa
     6121 aaagagcagt gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta
     6181 tgggcgcagc gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc
     6241 agcagcagaa caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag
     6301 tctggggcat caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc
     6361 aacagctcct ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt
     6421 ggaatgctag ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg
     6481 agtgggacag agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc
     6541 aaaaccagca agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt
     6601 ggaattggtt taacataaca aattggctgt ggtatataaa attattcata atgatagtag
     6661 gaggcttggt aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc
     6721 agggatattc accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc
     6781 ccgaaggaat agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga
     6841 acggatctcg acggtatcga tcacgagact agcctcgagc ggccgccccc ttcaccgagg
     6901 gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt agagagataa
     6961 ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga cgtagaaagt
     7021 aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac tatcatatgc
//
