LOCUS       sequence-347783-        7771 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Same as pLL3.3;U6(BstXI)-XhoI-chimeric RNA;PGK-Cre but XhoI stuffer 
            removed, expresses non-targeting gRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7771)
  AUTHORS   Murray CW, Brady JJ, Han M, Cai H, Tsai MK, Pierce SE, Cheng R, 
            Demeter J, Feldser DM, Jackson PK, Shackelford DB, Winslow MM
  TITLE     LKB1 drives stasis and C/EBP-mediated reprogramming to an alveolar 
            type II fate in lung cancer.
  JOURNAL   Nat Commun. 2022 Feb 28;13(1):1090. doi: 10.1038/s41467-022-28619-8.
  PUBMED    35228570
REFERENCE   2  (bases 1 to 7771)
  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: "2022-02-28"; volume: "13(1):1090. doi"; pages: " 
            10.1038/s41467-022-28619-8"
COMMENT     Sequence Label: hU6-sgNT1 (opti)
FEATURES             Location/Qualifiers
     source          1..7771
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(325..336)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(445..461)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(446..462)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             971..1151
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(1213..1801)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1293..1312)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1972..2763,2764..2832))
                     /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     2595..2614
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2833..2937)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3012..3031)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3203..3582
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3584..3782
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3737..3757
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             3800..3980
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4027..4152
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    4651..4884
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             5069..5113
                     /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    5411..5528
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5610..5858
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     5610..5630
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     5781..5800
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        6017..6516
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6052..6075)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     6449..6468
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             6520..6540
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6541..7569
                     /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="SNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKMLL
                     SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP
                     RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL
                     GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER
                     WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR
                     YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED
                     GD"
     misc_feature    join(7625..7771,1..442)
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7678..7698)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
ORIGIN
        1 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
       61 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
      121 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
      181 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
      241 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
      301 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
      361 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
      421 tccctttggg ccgcctcccc gcatcgatac cgtcgacctc gatcgagacc tagaaaaaca
      481 tggagcaatc acaagtagca atacagcagc taccaatgct gattgtgcct ggctagaagc
      541 acaagaggag gaggaggtgg gttttccagt cacacctcag gtacctttaa gaccaatgac
      601 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct
      661 aattcactcc caacgaagac aagatatcct tgatctgtgg atctaccaca cacaaggcta
      721 cttccctgat tggcagaact acacaccagg gccagggatc agatatccac tgacctttgg
      781 atggtgctac aagctagtac cagttgagca agagaaggta gaagaagcca atgaaggaga
      841 gaacacccgc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg agagagaagt
      901 attagagtgg aggtttgaca gccgcctagc atttcatcac atggcccgag agctgcatcc
      961 ggactgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     1021 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     1081 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     1141 aatctctagc agcatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     1201 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     1261 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     1321 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     1381 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     1441 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     1501 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     1561 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     1621 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     1681 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     1741 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     1801 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     1861 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     1921 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     1981 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     2041 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     2101 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     2161 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     2221 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     2281 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     2341 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     2401 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     2461 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     2521 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     2581 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     2641 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     2701 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     2761 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     2821 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     2881 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     2941 atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg
     3001 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
     3061 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
     3121 ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat
     3181 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     3241 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     3301 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     3361 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     3421 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     3481 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     3541 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     3601 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     3661 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     3721 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     3781 agcgcgtttt gcctgtactg ggtctctctg gttagaccag atctgagcct gggagctctc
     3841 tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt
     3901 agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc
     3961 agtgtggaaa atctctagca gtggcgcccg aacagggact tgaaagcgaa agggaaacca
     4021 gaggagctct ctcgacgcag gactcggctt gctgaagcgc gcacggcaag aggcgagggg
     4081 cggcgactgg tgagtacgcc aaaaattttg actagcggag gctagaagga gagagatggg
     4141 tgcgagagcg tcagtattaa gcgggggaga attagatcgc gatgggaaaa aattcggtta
     4201 aggccagggg gaaagaaaaa atataaatta aaacatatag tatgggcaag cagggagcta
     4261 gaacgattcg cagttaatcc tggcctgtta gaaacatcag aaggctgtag acaaatactg
     4321 ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt atataataca
     4381 gtagcaaccc tctattgtgt gcatcaaagg atagagataa aagacaccaa ggaagcttta
     4441 gacaagatag aggaagagca aaacaaaagt aagaccaccg cacagcaagc ggccggccgc
     4501 gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg aattatataa
     4561 atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt
     4621 ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc
     4681 agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt
     4741 gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct
     4801 gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag
     4861 atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac
     4921 cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca
     4981 cacgacctgg atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt
     5041 aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa
     5101 atgggcaagt ttgtggaatt ggtttaacat aacaaattgg ctgtggtata taaaattatt
     5161 cataatgata gtaggaggct tggtaggttt aagaatagtt tttgctgtac tttctatagt
     5221 gaatagagtt aggcagggat attcaccatt atcgtttcag acccacctcc caaccccgag
     5281 gggacccgac aggcccgaag gaatagaaga agaaggtgga gagagagaca gagacagatc
     5341 cattcgatta gtgaacggat cggcactgcg tgcgccaatt ctgcagacaa atggcagtat
     5401 tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag
     5461 tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc
     5521 aaaattttcg ggtttattac agggacagca gagatccagt ttggttagta ccgggcccgc
     5581 tctagccctg caggaactgt ttaaactgtg agggcctatt tcccatgatt ccttcatatt
     5641 tgcatatacg atacaaggct gttagagaga taattggaat taatttgact gtaaacacaa
     5701 agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt
     5761 taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt
     5821 tcttggcttt atatatcttg tggaaaggac gaaacaccgc gaggtattcg gctccgcggt
     5881 ttaagagcta tgctggaaac agcatagcaa gtttaaataa ggctagtccg ttatcaactt
     5941 gaaaaagtgg caccgagtcg gtgctttttt gtttggcgcg cctatggctt aattaacatg
     6001 gcggccgcat tctaccgggt aggggaggcg cttttcccaa ggcagtctgg agcatgcgct
     6061 ttagcagccc cgctgggcac ttggcgctac acaagtggcc tctggcctcg cacacattcc
     6121 acatccaccg gtaggcgcca accggctccg ttctttggtg gccccttcgc gccaccttct
     6181 actcctcccc tagtcaggaa gttccccccc gccccgcagc tcgcgtcgtg caggacgtga
     6241 caaatggaag tagcacgtct cactagtctc gtgcagatgg acagcaccgc tgagcaatgg
     6301 aagcgggtag gcctttgggg cagcggccaa tagcagcttt gctccttcgc tttctgggct
     6361 cagaggctgg gaaggggtgg gtccgggggc gggctcaggg gcgggctcag gggcggggcg
     6421 ggcgcccgaa ggtcctccgg aggcccggca ttctgcacgc ttcaaaagcg cacgtctgcc
     6481 gcgctgttct cctcttcctc atctccggga cccgggatgc ccaagaagaa gaggaaggtg
     6541 tccaatttac tgaccgtaca ccaaaatttg cctgcattac cggtcgatgc aacgagtgat
     6601 gaggttcgca agaacctgat ggacatgttc agggatcgcc aggcgttttc tgagcatacc
     6661 tggaaaatgc ttctgtccgt ttgccggtcg tgggcggcat ggtgcaagtt gaataaccgg
     6721 aaatggtttc ccgcagaacc tgaagatgtt cgcgattatc ttctatatct tcaggcgcgc
     6781 ggtctggcag taaaaactat ccagcaacat ttgggccagc taaacatgct tcatcgtcgg
     6841 tccgggctgc cacgaccaag tgacagcaat gctgtttcac tggttatgcg gcggatccga
     6901 aaagaaaacg ttgatgccgg tgaacgtgca aaacaggctc tagcgttcga acgcactgat
     6961 ttcgaccagg ttcgttcact catggaaaat agcgatcgct gccaggatat acgtaatctg
     7021 gcatttctgg ggattgctta taacaccctg ttacgtatag ccgaaattgc caggatcagg
     7081 gttaaagata tctcacgtac tgacggtggg agaatgttaa tccatattgg cagaacgaaa
     7141 acgctggtta gcaccgcagg tgtagagaag gcacttagcc tgggggtaac taaactggtc
     7201 gagcgatgga tttccgtctc tggtgtagct gatgatccga ataactacct gttttgccgg
     7261 gtcagaaaaa atggtgttgc cgcgccatct gccaccagcc agctatcaac tcgcgccctg
     7321 gaagggattt ttgaagcaac tcatcgattg atttacggcg ctaaggatga ctctggtcag
     7381 agatacctgg cctggtctgg acacagtgcc cgtgtcggag ccgcgcgaga tatggcccgc
     7441 gctggagttt caataccgga gatcatgcaa gctggtggct ggaccaatgt aaatattgtc
     7501 atgaactata tccgtaacct ggatagtgaa acaggggcaa tggtgcgcct gctggaagat
     7561 ggcgattagc catgtttaag ggttccggtt ccactaggta caattcgata tcaagcttat
     7621 cgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     7681 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     7741 ccgtatggct ttcattttct cctccttgta t
//
