LOCUS       sequence-318871-        7080 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SHIP1 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 Dec 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 SHIP1 KD2
FEATURES             Location/Qualifiers
     source          1..7080
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        80..328
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     80..100
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     251..270
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    421..538
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        587..1097
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     1016..1034
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             1119..1718
                     /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(1119..1138)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1615..1635
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1846..2079
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2151..2272
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2188..2207)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2242..2261
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2312..2447
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2467..2486)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2468..2486)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2496..2513)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2496..2512)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2505..2527)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2654..3109
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2741..2760)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2951..2972
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3135..3239
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3240..3308,3309..4100)
                     /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(3458..3477)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4271..4859
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4760..4779
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5013..5030
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5147..5168
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5183..5188,5189..5206,5207..5213)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5221..5237
                     /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     5226..5248
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5245..5261
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5245..5261
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5280..5300
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5282..5300
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5328..5554
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5555..5735
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5782..5907
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6400..6633
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6818..6862
                     /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"
ORIGIN
        1 agacagatcc attcgattag tgaacggatc tcgacggtat cgatcacgag actagcctcg
       61 agcggccgcc cccttcaccg agggcctatt tcccatgatt ccttcatatt tgcatatacg
      121 atacaaggct gttagagaga taattggaat taatttgact gtaaacacaa agatattagt
      181 acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt taaaattatg
      241 ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt tcttggcttt
      301 atatatcttg tggaaaggac gaaacaccgg agacgcggga tgattctgcc tcgaggcaga
      361 atcatcccgc gtcttttttg aattctcgac ctcgagacaa atggcagtat tcatccacaa
      421 ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat
      481 agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc aaaattttcg
      541 ggtttattac agggacagca gagatccact ttggccgcgg ctcgaggggg ttggggttgc
      601 gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg cgtggttccg
      661 ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt cgcagcgtca
      721 cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct ccgcccctaa
      781 gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa gccgcacgtc
      841 tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc cgaccgcgat
      901 gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc cgggaagggg
      961 cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc gcgcggtgtt
     1021 ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt gaccgaatca
     1081 ccgacctctc tccccagggg gatccaccgg agcttaccat gaccgagtac aagcccacgg
     1141 tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc gccgcgttcg
     1201 ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag cgggtcaccg
     1261 agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg
     1321 acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg ggggcggtgt
     1381 tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc gcgcagcaac
     1441 agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg
     1501 tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg ctccccggag
     1561 tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg ccccgcaacc
     1621 tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac
     1681 cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac gacccgcagc
     1741 gcccgaccga aaggagcgca cgaccccatg catcggtacc tttaagacca atgacttaca
     1801 aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc
     1861 actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg ttagaccaga
     1921 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
     1981 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
     2041 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca tgtcatctta
     2101 ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg aacttgttta
     2161 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     2221 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     2281 ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     2341 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     2401 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag ggacgtaccc
     2461 aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt
     2521 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     2581 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     2641 aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     2701 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     2761 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     2821 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     2881 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     2941 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     3001 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     3061 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt
     3121 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     3181 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     3241 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     3301 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     3361 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     3421 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     3481 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     3541 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     3601 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     3661 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     3721 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     3781 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     3841 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     3901 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     3961 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     4021 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     4081 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     4141 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     4201 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     4261 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     4321 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     4381 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     4441 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     4501 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     4561 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     4621 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     4681 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     4741 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     4801 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     4861 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     4921 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     4981 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     5041 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     5101 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
     5161 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
     5221 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg
     5281 caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgt agtcttatgc
     5341 aatactcttg tagtcttgca acatggtaac gatgagttag caacatgcct tacaaggaga
     5401 gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg ccttattagg
     5461 aaggcaacag acgggtctga catggattgg acgaaccact gaattgccgc attgcagaga
     5521 tattgtattt aagtgcctag ctcgatacat aaacgggtct ctctggttag accagatctg
     5581 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     5641 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     5701 cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa
     5761 gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg
     5821 gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag
     5881 aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg
     5941 gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg
     6001 gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc
     6061 tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga
     6121 tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac
     6181 accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag
     6241 caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt ggagaagtga
     6301 attatataaa tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa
     6361 gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt
     6421 cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg tacaggccag
     6481 acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca
     6541 acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa gaatcctggc
     6601 tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact
     6661 catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat
     6721 ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca caagcttaat
     6781 acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag aattattgga
     6841 attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc tgtggtatat
     6901 aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt ttgctgtact
     6961 ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga cccacctccc
     7021 aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag agagagacag
//
