LOCUS       sequence-363747-        7748 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses a Arid1a-targeting gRNA and Cre-recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7748)
  AUTHORS   Cai H, Chew SK, Li C, Tsai MK, Andrejka L, Murray CW, Hughes NW, 
            Shuldiner EG, Ashkin EL, Tang R, Hung KL, Chen LC, Lee SYC, Yousefi 
            M, Lin WY, Kunder CA, Cong L, McFarland CD, Petrov DA, Swanton C, 
            Winslow MM
  TITLE     A functional taxonomy of tumor suppression in oncogenic KRAS-driven 
            lung cancer.
  JOURNAL   Cancer Discov. 2021 Feb 19. pii: 2159-8290.CD-20-1325. doi: 
            10.1158/2159-8290.CD-20-1325.
  PUBMED    33608386
REFERENCE   2  (bases 1 to 7748)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cancer 
            Discov. 2021 Feb 19. pii: 2159-8290.CD-20-1325. doi: 
            10.1158/2159-8290.CD-20-1325."
COMMENT     Sequence Label: Lenti-sgArid1a#2/Cre
FEATURES             Location/Qualifiers
     source          1..7748
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(6..25)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        197..576
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        578..776
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     731..751
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             794..974
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1021..1146
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1643..1876
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2061..2105
                     /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"
     misc_feature    2403..2520
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2602..2850
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2602..2622
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2773..2792
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2872..2947
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3000..3487
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3035..3058)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3420..3439
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3491..3511
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3512..4540
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="SNLLTVHQNLPALPVDATSDEVSKNLMDMFRDRQAFSEHTWKMLL
                     SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP
                     RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL
                     GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER
                     WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR
                     YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED
                     GD"
     misc_feature    4596..5184
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4649..4669)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5067..5078)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5187..5203)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5188..5204)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5713..5893
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5955..6543)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6035..6054)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6714..7505,7506..7574))
                     /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     7337..7356
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7575..7679)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg
       61 ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc
      121 gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg
      181 gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg
      241 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      301 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      361 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      421 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      481 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      541 cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc
      601 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
      661 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
      721 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagcgcg
      781 ttttgcctgt actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta
      841 actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg
      901 tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg
      961 gaaaatctct agcagtggcg cccgaacagg gacttgaaag cgaaagggaa accagaggag
     1021 ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga
     1081 ctggtgagta cgccaaaaat tttgactagc ggaggctaga aggagagaga tgggtgcgag
     1141 agcgtcagta ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg gttaaggcca
     1201 gggggaaaga aaaaatataa attaaaacat atagtatggg caagcaggga gctagaacga
     1261 ttcgcagtta atcctggcct gttagaaaca tcagaaggct gtagacaaat actgggacag
     1321 ctacaaccat cccttcagac aggatcagaa gaacttagat cattatataa tacagtagca
     1381 accctctatt gtgtgcatca aaggatagag ataaaagaca ccaaggaagc tttagacaag
     1441 atagaggaag agcaaaacaa aagtaagacc accgcacagc aagcggccgg ccgctgatct
     1501 tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat aaatataaag
     1561 tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga
     1621 gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa
     1681 gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta
     1741 tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac
     1801 tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa
     1861 aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg
     1921 tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct
     1981 ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag
     2041 aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa
     2101 gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga
     2161 tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag
     2221 ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg
     2281 acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat
     2341 tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac aaatggcagt attcatccac
     2401 aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata
     2461 atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt
     2521 cgggtttatt acagggacag cagagatcca gtttggttag taccgggccc gctctagccc
     2581 tgcaggaact gtttaaactg tgagggccta tttcccatga ttccttcata tttgcatata
     2641 cgatacaagg ctgttagaga gataattgga attaatttga ctgtaaacac aaagatatta
     2701 gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta
     2761 tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct
     2821 ttatatatct tgtggaaagg accaaacacc gcctgctggc catacgcact ggttttagag
     2881 ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag
     2941 tcggtgcttt tttgtttggc gcgcctatgg cttaattaac atggcggccg cattctaccg
     3001 ggtaggggag gcgcttttcc caaggcagtc tggagcatgc gctttagcag ccccgctggg
     3061 cacttggcgc tacacaagtg gcctctggcc tcgcacacat tccacatcca ccggtaggcg
     3121 ccaaccggct ccgttctttg gtggcccctt cgcgccacct tctactcctc ccctagtcag
     3181 gaagttcccc cccgccccgc agctcgcgtc gtgcaggacg tgacaaatgg aagtagcacg
     3241 tctcactagt ctcgtgcaga tggacagcac cgctgagcaa tggaagcggg taggcctttg
     3301 gggcagcggc caatagcagc tttgctcctt cgctttctgg gctcagaggc tgggaagggg
     3361 tgggtccggg ggcgggctca ggggcggggc gggcgcccga aggtcctccg gaggcccggc
     3421 attctgcacg cttcaaaagc gcacgtctgc cgcgctgttc tcctcttcct catctccggg
     3481 acccgggatg cccaagaaga agaggaaggt gtccaattta ctgaccgtac accaaaattt
     3541 gcctgcatta ccggtcgatg caacgagtga tgaggttagc aagaacctga tggacatgtt
     3601 cagggatcgc caggcgtttt ctgagcatac ctggaaaatg cttctgtccg tttgccggtc
     3661 gtgggcggca tggtgcaagt tgaataaccg gaaatggttt cccgcagaac ctgaagatgt
     3721 tcgcgattat cttctatatc ttcaggcgcg cggtctggca gtaaaaacta tccagcaaca
     3781 tttgggccag ctaaacatgc ttcatcgtcg gtccgggctg ccacgaccaa gtgacagcaa
     3841 tgctgtttca ctggttatgc ggcggatccg aaaagaaaac gttgatgccg gtgaacgtgc
     3901 aaaacaggct ctagcgttcg aacgcactga tttcgaccag gttcgttcac tcatggaaaa
     3961 tagcgatcgc tgccaggata tacgtaatct ggcatttctg gggattgctt ataacaccct
     4021 gttacgtata gccgaaattg ccaggatcag ggttaaagat atctcacgta ctgacggtgg
     4081 gagaatgtta atccatattg gcagaacgaa aacgctggtt agcaccgcag gtgtagagaa
     4141 ggcacttagc ctgggggtaa ctaaactggt cgagcgatgg atttccgtct ctggtgtagc
     4201 tgatgatccg aataactacc tgttttgccg ggtcagaaaa aatggtgttg ccgcgccatc
     4261 tgccaccagc cagctatcaa ctcgcgccct ggaagggatt tttgaagcaa ctcatcgatt
     4321 gatttacggc gctaaggatg actctggtca gagatacctg gcctggtctg gacacagtgc
     4381 ccgtgtcgga gccgcgcgag atatggcccg cgctggagtt tcaataccgg agatcatgca
     4441 agctggtggc tggaccaatg taaatattgt catgaactat atccgtaacc tggatagtga
     4501 aacaggggca atggtgcgcc tgctggaaga tggcgattag ccatgtttaa gggttccggt
     4561 tccactaggt acaattcgat atcaagctta tcgataatca acctctggat tacaaaattt
     4621 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     4681 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     4741 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     4801 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     4861 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     4921 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     4981 tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc
     5041 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     5101 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     5161 tctccctttg ggccgcctcc ccgcatcgat accgtcgacc tcgatcgaga cctagaaaaa
     5221 catggagcaa tcacaagtag caatacagca gctaccaatg ctgattgtgc ctggctagaa
     5281 gcacaagagg aggaggaggt gggttttcca gtcacacctc aggtaccttt aagaccaatg
     5341 acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg
     5401 ctaattcact cccaacgaag acaagatatc cttgatctgt ggatctacca cacacaaggc
     5461 tacttccctg attggcagaa ctacacacca gggccaggga tcagatatcc actgaccttt
     5521 ggatggtgct acaagctagt accagttgag caagagaagg tagaagaagc caatgaagga
     5581 gagaacaccc gcttgttaca ccctgtgagc ctgcatggga tggatgaccc ggagagagaa
     5641 gtattagagt ggaggtttga cagccgccta gcatttcatc acatggcccg agagctgcat
     5701 ccggactgta ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa
     5761 ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt
     5821 gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg
     5881 aaaatctcta gcagcatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     5941 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     6001 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     6061 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     6121 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     6181 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     6241 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     6301 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     6361 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     6421 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     6481 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     6541 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     6601 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     6661 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     6721 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     6781 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     6841 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     6901 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     6961 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     7021 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     7081 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     7141 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     7201 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     7261 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     7321 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     7381 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     7441 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     7501 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     7561 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     7621 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     7681 acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat
     7741 ggtgcact
//
