LOCUS       sequence-261525-        8701 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Forced expression of NKX6.1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8701)
  TITLE     Lian Lab CRISPR plasmids
REFERENCE   2  (bases 1 to 8701)
  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"
COMMENT     Sequence Label: pYJ09-lenti-EF1a-NKX6.1-puro
FEATURES             Location/Qualifiers
     source          1..8701
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             117..750
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    797..922
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1415..1648
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1833..1877
                     /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    2164..2281
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2410..3588
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2641..3579
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3536..3556
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     misc_feature    join(4817..5391,5392..5394,5395..5403)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4984..5001)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     5211..5230
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5423..6022
                     /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(5423..5442)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5919..5939
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             6225..6458
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(6609..6626)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6615..6839
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     regulatory      6882..6891
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        6935..7039
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7040..7108,7109..7900)
                     /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(7258..7277)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8071..8659
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8560..8579
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cggaccgcca ctgccaatta cctgtggttt catttactct aaacctgtga ttcctctgaa
       61 ttattttcat tttaaagaaa ttgtatttgt taaatatgta ctacaaactt agtagttgga
      121 agggctaatt cactcccaaa gaagacaaga tatccttgat ctgtggatct accacacaca
      181 aggctacttc cctgattagc agaactacac accagggcca ggggtcagat atccactgac
      241 ctttggatgg tgctacaagc tagtaccagt tgagccagat aaggtagaag aggccaataa
      301 aggagagaac accagcttgt tacaccctgt gagcctgcat gggatggatg acccggagag
      361 agaagtgtta gagtggaggt ttgacagccg cctagcattt catcacgtgg cccgagagct
      421 gcatccggag tacttcaaga actgctgata tcgagcttgc tacaagggac tttccgctgg
      481 ggactttcca gggaggcgtg gcctgggcgg gactggggag tggcgagccc tcagatcctg
      541 catataagca gctgcttttt gcctgtactg ggtctctctg gttagaccag atctgagcct
      601 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
      661 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
      721 ccttttagtc agtgtggaaa atctctagca gtggcgcccg aacagggact tgaaagcgaa
      781 agggaaacca gaggagctct ctcgacgcag gactcggctt gctgaagcgc gcacggcaag
      841 aggcgagggg cggcgactgg tgagtacgcc aaaaattttg actagcggag gctagaagga
      901 gagagatggg tgcgagagcg tcagtattaa gcgggggaga attagatcgc gatgggaaaa
      961 aattcggtta aggccagggg gaaagaaaaa atataaatta aaacatatag tatgggcaag
     1021 cagggagcta gaacgattcg cagttaatcc tggcctgtta gaaacatcag aaggctgtag
     1081 acaaatactg ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt
     1141 atataataca gtagcaaccc tctattgtgt gcatcaaagg atagagataa aagacaccaa
     1201 ggaagcttta gacaagatag aggaagagca aaacaaaagt aagaccaccg cacagcaagc
     1261 ggccgctgat cttcagacct ggaggaggag atatgaggga caattggaga agtgaattat
     1321 ataaatataa agtagtaaaa attgaaccat taggagtagc acccaccaag gcaaagagaa
     1381 gagtggtgca gagagaaaaa agagcagtgg gaataggagc tttgttcctt gggttcttgg
     1441 gagcagcagg aagcactatg ggcgcagcgt caatgacgct gacggtacag gccagacaat
     1501 tattgtctgg tatagtgcag cagcagaaca atttgctgag ggctattgag gcgcaacagc
     1561 atctgttgca actcacagtc tggggcatca agcagctcca ggcaagaatc ctggctgtgg
     1621 aaagatacct aaaggatcaa cagctcctgg ggatttgggg ttgctctgga aaactcattt
     1681 gcaccactgc tgtgccttgg aatgctagtt ggagtaataa atctctggaa cagatttgga
     1741 atcacacgac ctggatggag tgggacagag aaattaacaa ttacacaagc ttaatacact
     1801 ccttaattga agaatcgcaa aaccagcaag aaaagaatga acaagaatta ttggaattag
     1861 ataaatgggc aagtttgtgg aattggttta acataacaaa ttggctgtgg tatataaaat
     1921 tattcataat gatagtagga ggcttggtag gtttaagaat agtttttgct gtactttcta
     1981 tagtgaatag agttaggcag ggatattcac cattatcgtt tcagacccac ctcccaaccc
     2041 cgaggggacc cgacaggccc gaaggaatag aagaagaagg tggagagaga gacagagaca
     2101 gatccattcg attagtgaac ggatctcgac ggtatcgcca caaatggcag tattcatcca
     2161 caattttaaa agaaaggggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     2221 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     2281 tcgggtttat tacagggaca gcagagatcc actttggatc gataagcttt gcaaagatgg
     2341 ataaagtttt aaacagagag gaatctttgc agctaatgga ccttctaggt cttgaaagga
     2401 gtgggaattg gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag
     2461 aagttggggg gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac
     2521 tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat
     2581 ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag
     2641 gtaagtgccg tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg
     2701 ccttgaatta cttccacctg gctgcagtac gtgattcttg atcccgagct tcgggttgga
     2761 agtgggtggg agagttcgag gccttgcgct taaggagccc cttcgcctcg tgcttgagtt
     2821 gaggcctggc ctgggcgctg gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt
     2881 ctcgctgctt tcgataagtc tctagccatt taaaattttt gatgacctgc tgcgacgctt
     2941 tttttctggc aagatagtct tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt
     3001 tttggggccg cgggcggcga cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg
     3061 ggcctgcgag cgcggccacc gagaatcgga cgggggtagt ctcaagctgg ccggcctgct
     3121 ctggtgcctg gcctcgcgcc gccgtgtatc gccccgccct gggcggcaag gctggcccgg
     3181 tcggcaccag ttgcgtgagc ggaaagatgg ccgcttcccg gccctgctgc agggagctca
     3241 aaatggagga cgcggcgctc gggagagcgg gcgggtgagt cacccacaca aaggaaaagg
     3301 gcctttccgt cctcagccgt cgcttcatgt gactccacgg agtaccgggc gccgtccagg
     3361 cacctcgatt agttctcgag cttttggagt acgtcgtctt taggttgggg ggaggggttt
     3421 tatgcgatgg agtttcccca cactgagtgg gtggagactg aagttaggcc agcttggcac
     3481 ttgatgtaat tctccttgga atttgccctt tttgagtttg gatcttggtt cattctcaag
     3541 cctcagacag tggttcaaag tttttttctt ccatttcagg tgtcgtgagg aattgatcct
     3601 tcgaactagt gccaccatgt tagcggtggg ggcaatggag ggcacccggc agagcgcatt
     3661 cctgctcagc agccctcccc tggccgccct gcacagcatg gccgagatga agaccccgct
     3721 gtaccctgcc gcgtatcccc cgctgcctgc cggccccccc tcctcctcgt cctcgtcgtc
     3781 gtcctcctcg tcgccctccc cgcctctggg cacccacaac ccaggcggcc tgaagccccc
     3841 ggccacgggg gggctctcat ccctcggcag ccccccgcag cagctctcgg ccgccacccc
     3901 acacggcatc aacgatatcc tgagccggcc ctccatgccc gtggcctcgg gggccgccct
     3961 gccctccgcc tcgccctccg gttcctcctc ctcctcttcc tcgtccgcct ctgcctcctc
     4021 cgcctctgcc gccgccgcgg ctgctgccgc ggccgcagcc gccgcctcat ccccggcggg
     4081 gctgctggcc ggactgccac gctttagcag cctgagcccg ccgccgccgc cgcccgggct
     4141 ctacttcagc cccagcgccg cggccgtggc cgccgtgggc cggtacccca agccgctggc
     4201 tgagctgcct ggccggacgc ccatcttctg gcccggagtg atgcagagcc cgccctggag
     4261 ggacgcacgc ctggcctgta cccctcatca aggatccatt ttgttggaca aagacgggaa
     4321 gagaaaacac acgagaccca ctttttccgg acagcagatc ttcgccctgg agaagacttt
     4381 cgaacaaaca aaatacttgg cggggcccga gagggctcgt ttggcctatt cgttggggat
     4441 gacagagagt caggtcaagg tctggttcca gaaccgccgg accaagtgga ggaagaagca
     4501 cgctgccgag atggccacgg ccaagaagaa gcaggactcg gagacagagc gcctcaaggg
     4561 ggcctcggag aacgaggaag aggacgacga ctacaataag cctctggatc ccaactcgga
     4621 cgacgagaaa atcacgcagc tgttgaagaa gcacaagtcc agcagcggcg gcggcggcgg
     4681 cctcctactg cacgcgtccg agccggagag ctcatcctga gaattcgaag cttgagctcg
     4741 agatctgagt ccggtagcgc tagttcgaag gatccgcatg catctagggc ggccaattcc
     4801 gcccctctcc cccccccccc tctccctccc ccccccctaa cgttactggc cgaagccgct
     4861 tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg ccgtcttttg
     4921 gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct aggggtcttt
     4981 cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg
     5041 aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg aaccccccac
     5101 ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct gcaaaggcgg
     5161 cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa tggctctcct
     5221 caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt atgggatctg
     5281 atctggggcc tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa aaacgtctag
     5341 gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgatgataa gcttgccaca
     5401 acccacaagg agacgacctt ccatgaccga gtacaagccc acggtgcgcc tcgccacccg
     5461 cgacgacgtc ccccgggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac
     5521 gcgccacacc gtcgacccgg accgccacat cgagcgggtc accgagctgc aagaactctt
     5581 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt
     5641 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc
     5701 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct
     5761 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga
     5821 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg
     5881 cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg
     5941 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat
     6001 gacccgcaag cccggtgcct gataataggc ggccgctcga gacctagaaa aacatggagc
     6061 aatcacaagt agcaatacag cagctaccaa tgctgattgt gcctggctag aagcacaaga
     6121 ggaggaggag gtgggttttc cagtcacacc tcaggtacct ttaagaccaa tgacttacaa
     6181 ggcagctgta gatcttagcc actttttaaa agaaaagggg ggactggaag ggctaattca
     6241 ctcccaacga agacaagatc tgctttttgc ttgtactggg tctctctggt tagaccagat
     6301 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
     6361 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
     6421 cctcagaccc ttttagtcag tgtggaaaat ctctagcagt agtagttcat gtcatcttat
     6481 tattcagtat ttataacttg caaagaaatg aatatcagag agtgagaggc cttgacatta
     6541 taatagattt agcaggaatt gaactaggag tggagcacac aggcaaagtt ctagagctcg
     6601 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     6661 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     6721 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     6781 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     6841 ttctgaggcg gaaagaacca gctgggggcg cgcccctcga ggccgccatg gtcatagctg
     6901 tttgacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     6961 ctaaatacat tcaaatatgt atccgctcac gagacaataa ccctgataaa tgcttcaata
     7021 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     7081 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     7141 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     7201 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     7261 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     7321 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     7381 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     7441 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     7501 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     7561 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     7621 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     7681 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     7741 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     7801 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     7861 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     7921 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     7981 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     8041 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     8101 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     8161 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     8221 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     8281 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     8341 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     8401 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     8461 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     8521 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     8581 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     8641 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttct ggccttttgc
     8701 a
//
