LOCUS       sequence-352701-        7769 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 7769)
  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 7769)
  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-sgNT3 (opti)
FEATURES             Location/Qualifiers
     source          1..7769
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(401..412)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(521..537)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(522..538)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             1047..1227
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(1289..1877)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1369..1388)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2048..2839,2840..2908))
                     /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     2671..2690
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2909..3013)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3088..3107)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3279..3658
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3660..3858
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3813..3833
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             3876..4056
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4103..4228
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    4725..4958
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             5143..5187
                     /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    5485..5602
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5684..5932
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     5684..5704
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     5855..5874
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        6091..6590
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6126..6149)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     6523..6542
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             6594..6614
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6615..7643
                     /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(7699..7769,1..518)
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(join(7752..7769,1..3))
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
ORIGIN
        1 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat
       61 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt
      121 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat
      181 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc
      241 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
      301 caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc
      361 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga
      421 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc
      481 tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgtc gacctcgatc
      541 gagacctaga aaaacatgga gcaatcacaa gtagcaatac agcagctacc aatgctgatt
      601 gtgcctggct agaagcacaa gaggaggagg aggtgggttt tccagtcaca cctcaggtac
      661 ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta aaagaaaagg
      721 ggggactgga agggctaatt cactcccaac gaagacaaga tatccttgat ctgtggatct
      781 accacacaca aggctacttc cctgattggc agaactacac accagggcca gggatcagat
      841 atccactgac ctttggatgg tgctacaagc tagtaccagt tgagcaagag aaggtagaag
      901 aagccaatga aggagagaac acccgcttgt tacaccctgt gagcctgcat gggatggatg
      961 acccggagag agaagtatta gagtggaggt ttgacagccg cctagcattt catcacatgg
     1021 cccgagagct gcatccggac tgtactgggt ctctctggtt agaccagatc tgagcctggg
     1081 agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc
     1141 ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct
     1201 tttagtcagt gtggaaaatc tctagcagca tgtgagcaaa aggccagcaa aaggccagga
     1261 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     1321 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     1381 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     1441 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     1501 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     1561 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     1621 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     1681 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     1741 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     1801 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     1861 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     1921 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     1981 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     2041 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     2101 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     2161 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     2221 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     2281 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     2341 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     2401 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     2461 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     2521 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     2581 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     2641 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     2701 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     2761 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     2821 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     2881 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     2941 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     3001 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     3061 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     3121 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     3181 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt
     3241 tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta
     3301 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
     3361 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
     3421 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
     3481 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
     3541 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
     3601 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
     3661 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
     3721 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
     3781 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
     3841 ggaggtctat ataagcagcg cgttttgcct gtactgggtc tctctggtta gaccagatct
     3901 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     3961 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     4021 tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa
     4081 agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac
     4141 ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta
     4201 gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg
     4261 ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg
     4321 ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg
     4381 ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag aagaacttag
     4441 atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga
     4501 caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca
     4561 gcaagcggcc ggccgctgat cttcagacct ggaggaggag atatgaggga caattggaga
     4621 agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc acccaccaag
     4681 gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc tttgttcctt
     4741 gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct gacggtacag
     4801 gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag ggctattgag
     4861 gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca ggcaagaatc
     4921 ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg ttgctctgga
     4981 aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa atctctggaa
     5041 cagatttgga atcacacgac ctggatggag tgggacagag aaattaacaa ttacacaagc
     5101 ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga acaagaatta
     5161 ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa ttggctgtgg
     5221 tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat agtttttgct
     5281 gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt tcagacccac
     5341 ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg tggagagaga
     5401 gacagagaca gatccattcg attagtgaac ggatcggcac tgcgtgcgcc aattctgcag
     5461 acaaatggca gtattcatcc acaattttaa aagaaaaggg gggattgggg ggtacagtgc
     5521 aggggaaaga atagtagaca taatagcaac agacatacaa actaaagaat tacaaaaaca
     5581 aattacaaaa attcaaaatt ttcgggttta ttacagggac agcagagatc cagtttggtt
     5641 agtaccgggc ccgctctagc cctgcaggaa ctgtttaaac tgtgagggcc tatttcccat
     5701 gattccttca tatttgcata tacgatacaa ggctgttaga gagataattg gaattaattt
     5761 gactgtaaac acaaagatat tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg
     5821 gtagtttgca gttttaaaat tatgttttaa aatggactat catatgctta ccgtaacttg
     5881 aaagtatttc gatttcttgg ctttatatat cttgtggaaa ggaccaaaca ccgactgaaa
     5941 tccaaggact gtgtttaaga gctatgctgg aaacagcata gcaagtttaa ataaggctag
     6001 tccgttatca acttgaaaaa gtggcaccga gtcggtgctt ttttgtttgg cgcgcctatg
     6061 gcttaattaa catggcggcc gcattctacc gggtagggga ggcgcttttc ccaaggcagt
     6121 ctggagcatg cgctttagca gccccgctgg gcacttggcg ctacacaagt ggcctctggc
     6181 ctcgcacaca ttccacatcc accggtaggc gccaaccggc tccgttcttt ggtggcccct
     6241 tcgcgccacc ttctactcct cccctagtca ggaagttccc ccccgccccg cagctcgcgt
     6301 cgtgcaggac gtgacaaatg gaagtagcac gtctcactag tctcgtgcag atggacagca
     6361 ccgctgagca atggaagcgg gtaggccttt ggggcagcgg ccaatagcag ctttgctcct
     6421 tcgctttctg ggctcagagg ctgggaaggg gtgggtccgg gggcgggctc aggggcgggc
     6481 tcaggggcgg ggcgggcgcc cgaaggtcct ccggaggccc ggcattctgc acgcttcaaa
     6541 agcgcacgtc tgccgcgctg ttctcctctt cctcatctcc gggacccggg atgcccaaga
     6601 agaagaggaa ggtgtccaat ttactgaccg tacaccaaaa tttgcctgca ttaccggtcg
     6661 atgcaacgag tgatgaggtt cgcaagaacc tgatggacat gttcagggat cgccaggcgt
     6721 tttctgagca tacctggaaa atgcttctgt ccgtttgccg gtcgtgggcg gcatggtgca
     6781 agttgaataa ccggaaatgg tttcccgcag aacctgaaga tgttcgcgat tatcttctat
     6841 atcttcaggc gcgcggtctg gcagtaaaaa ctatccagca acatttgggc cagctaaaca
     6901 tgcttcatcg tcggtccggg ctgccacgac caagtgacag caatgctgtt tcactggtta
     6961 tgcggcggat ccgaaaagaa aacgttgatg ccggtgaacg tgcaaaacag gctctagcgt
     7021 tcgaacgcac tgatttcgac caggttcgtt cactcatgga aaatagcgat cgctgccagg
     7081 atatacgtaa tctggcattt ctggggattg cttataacac cctgttacgt atagccgaaa
     7141 ttgccaggat cagggttaaa gatatctcac gtactgacgg tgggagaatg ttaatccata
     7201 ttggcagaac gaaaacgctg gttagcaccg caggtgtaga gaaggcactt agcctggggg
     7261 taactaaact ggtcgagcga tggatttccg tctctggtgt agctgatgat ccgaataact
     7321 acctgttttg ccgggtcaga aaaaatggtg ttgccgcgcc atctgccacc agccagctat
     7381 caactcgcgc cctggaaggg atttttgaag caactcatcg attgatttac ggcgctaagg
     7441 atgactctgg tcagagatac ctggcctggt ctggacacag tgcccgtgtc ggagccgcgc
     7501 gagatatggc ccgcgctgga gtttcaatac cggagatcat gcaagctggt ggctggacca
     7561 atgtaaatat tgtcatgaac tatatccgta acctggatag tgaaacaggg gcaatggtgc
     7621 gcctgctgga agatggcgat tagccatgtt taagggttcc ggttccacta ggtacaattc
     7681 gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     7741 attcttaact atgttgctcc ttttacgct
//
