LOCUS       sequence-348093-        7057 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  knockout mouse Tpi1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7057)
  AUTHORS   Wu L, Hollinshead KER, Hao Y, Au C, Kroehling L, Ng C, Lin WY, Li D,
            Silva HM, Shin J, Lafaille JJ, Possemato R, Pacold ME, 
            Papagiannakopoulos T, Kimmelman AC, Satija R, Littman DR
  TITLE     Niche-Selective Inhibition of Pathogenic Th17 Cells by Targeting 
            Metabolic Redundancy.
  JOURNAL   Cell. 2020 Aug 6;182(3):641-654.e20. doi: 
            10.1016/j.cell.2020.06.014. Epub 2020 Jul 1.
  PUBMED    32615085
REFERENCE   2  (bases 1 to 7057)
  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"; doi: 
            "10.1016/j.cell.2020.06.014"; journalName: "Cell"; date: "2020-08-6-
            6"; volume: "182"; issue: "3"; pages: "641-654.e20"
COMMENT     Sequence Label: pSIN-Tpi1
FEATURES             Location/Qualifiers
     source          1..7057
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     181..201
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     337..359
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     LTR             462..887
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     CDS             complement(1008..1808)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="GPMIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQG
                     RPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEE
                     HQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQD
                     IALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1119..1138)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1729..1748
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(1803..1859)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(1869..2354)
                     /codon_start=1
                     /product="mouse GPI-anchored cell surface glycoprotein of 
                     the immunoglobulin superfamily"
                     /label=Thy1.1
                     /note="This allele of Thy1 is also known as CD90.1."
                     /translation="MNPAISVALLLSVLQVSRGQKVTSLTACLVNQNLRLDCRHENNTK
                     DNSIQHEFSLTREKRKHVLSGTLGIPEHTYRSRVTLSNQPYIKVLTLANFTTKDEGDYF
                     CELRVSGANPMSSNKSISVYRDKLVKCGGISLLVQNTSWMLLLLLSLSLLQALDFISL"
     promoter        complement(2374..2585)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     misc_RNA        complement(2692..2767)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(2789..3037)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(2847..2866)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(3017..3037)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     CDS             complement(3084..3499)
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(3100..3116)
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     complement(3132..3154)
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     misc_feature    3700..3899
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     LTR             3962..4137
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     promoter        complement(4138..4341)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4167..4187)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4342..4721
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(4821..4839)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4907..5011
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5012..5080,5081..5872)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5230..5249)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6043..6631
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6532..6551
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6785..6802
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(6883..6902)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        complement(join(6924..7057,1..196))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6938..7057,1..16)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gagttagggg cgggatgggc ggagttaggg gcgggactat ggttgctgac taattgagat
       61 gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac ctggttgctg
      121 actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac
      181 accctaactg acacacattc cacagccgga tctacactcc gctatcgcta cgtgactggg
      241 tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
      301 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
      361 tttcaccgtc atcaccgaaa cgcgcgaggc agccagctta cctcccggtg gtgggtcggt
      421 ggtccctggg caggggtctc ccgatcccgg acgagccccc aaatgaaaga cccccgctga
      481 cgggtagtca atcactcaga ggagaccctc ccaaggaaca gcgagaccac aagtcggatg
      541 caactgcaag agggtttatt ggatacacgg gtacccgggc gactcagtca atcggaggac
      601 tggcgccccg agtgaggggt tgtgggctct tttattgagc tcggggagca gaagcgcgcg
      661 aacagaagcg agaagcgaac tgattggtta gttcaaataa ggcacagggt catttcaggt
      721 ccttggggca ccctggaaac atctgatggt tctctagggt cgaccctgtt ccatctgttc
      781 ctgaccttga tctgaacttc tctattctca gttatgtatt tttccatgcc ttgcaaaatg
      841 gcgttactta agctagcttg ccaaacctac aggtggggtc tttcattccc ccctttttct
      901 ggagactaaa taaaatcttt tattttatct atggctcgta ctctataggc ttcagctggt
      961 gatattgttg agtcaaaact agagcctgga ccactgatga tatctaatca gaagaactcg
     1021 tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg
     1081 aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt agccaacgct
     1141 atgtcctgat agcggtccgc cacacccagc cggccacagt cgatgaatcc agaaaagcgg
     1201 ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac gagatcctcg
     1261 ccgtcgggca tgcgcgcctt gagcctggcg aacagttcgg ctggcgcgag cccctgatgc
     1321 tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg tgctcgctcg
     1381 atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt atgcagccgc
     1441 cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga tgacaggaga
     1501 tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt gacaacgtcg
     1561 agcacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc tgcctcgtcc
     1621 tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc
     1681 gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc ccagtcatag
     1741 ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc ttgttcaatc
     1801 atggggccgg ggttctcctc cacgtcgccg gcctgcttca gcaggctgaa gttggtggcg
     1861 ccgctgccca gagaaatgaa gtccagggct tggaggaggg agagggaaag cagcagcagc
     1921 agcatccagg atgtgttctg aaccagcagg cttatgccgc cacacttgac cagtttgtct
     1981 ctatacacac tgatactttt attggagctc atgggattcg cgcccgagac tcgaagctca
     2041 caaaagtagt cgccctcatc cttggtggtg aagttggcta gggtaaggac cttgatatag
     2101 ggctggttgg agagggtgac gcgggagcgg tacgtgtgct cgggtatccc gagggtgcct
     2161 gagagcacgt gcttcctctt ctctcgggtc aggctgaact catgctggat ggagttatcc
     2221 ttggtgttat tctcatggcg gcagtccagg cgaaggtttt ggttcaccag gcaggctgtc
     2281 aggctggtca ccttctgccc tcgggacacc tgcaagactg agagcaggag agcgacgctg
     2341 atggctgggt tcatggtggc cgcgtcacga cacctgtgtt ctggcggcaa acccgttgcg
     2401 aaaaagaacg ttcacggcga ctactgcact tatatacggt tctcccccac cctcgggaaa
     2461 aaggcggagc cagtacacga catcactttc ccagtttacc ccgcgccacc ttctctaggc
     2521 accggttcaa ttgccgaccc ctccccccaa cttctcgggg actgtgggcg atgtgcgctc
     2581 tgcccactga cgggcaccgg agccactcga ggggccggga gcatgcgacg tcgggcccaa
     2641 ttcgccgaat tcccggcccg gcaccttgtc tcaagatcta aattcaaaaa agcaccgact
     2701 cggtgccact ttttcaagtt gataacggac tagccttatt ttaacttgct atttctagct
     2761 ctaaaacctt acatcgactt tgccagacgg tgtttcgtcc tttccacaag atatataaag
     2821 ccaagaaatc gaaatacttt caagttacgg taagcatatg atagtccatt ttaaaacata
     2881 attttaaaac tgcaaactac ccaagaaatt attactttct acgtcacgta ttttgtacta
     2941 atatctttgt gtttacagtc aaattaattc taattatctc tctaacagcc ttgtatcgta
     3001 tatgcaaata tgaaggaatc atgggaaata ggccctctct agatctagaa tcgatagatc
     3061 gttaattaaa gatcttcaat tccggcgcct agagaaggag tgagggctgg ataaagggag
     3121 gatcgaggcg gggtcgaacg aggaggttca agggggagag acggggcgga tggaggaaga
     3181 ggaggcggag gcttagggtg tacaaagggc ttgacccagg gaggggggtc aaaagccaag
     3241 gcttcccagg tcacgatgta ggggacctgg tctgggtgtc catgcgggcc aggtgaaaag
     3301 accttgatct taacctgggt gatgaggtct cggttaaagg tgccgtctcg cggccatccg
     3361 acgttaaagg ttggccattc tgcagagcag aaggtaacca acgtctcttc ttgacatcta
     3421 ccgactggtt gtgagcgatc cgctcgacat ctttccagtg acctaaggtc aaacttaagg
     3481 gagtggtaac agtctggccc taattttcag acaaatacag aaacacagtc agacagagac
     3541 aacacagaac gatgctgcag cgctgcagca gacaagacgc gcggcttcgg ttccaaaccg
     3601 aaagcaaaaa ttcagacgga ggcgggaact gttttaggtt ctcgtctcct accagaacca
     3661 catatcctga cggggtcgga ttccacatcg actcccttcc tcaggtcggg ccacaaaaac
     3721 ggcccccaaa gtccctggga cgtctcccag ggttgcggcc gggtgttcag aactcgtcag
     3781 ttccaccacg ggtccgccag atacagagct agttagctaa ctagtacaga cgcaggcgca
     3841 taacatcaaa catagacact agacaatcgg acagacacag ataagttgct ggccagctta
     3901 cctcccggtg gtgggtcggt ggtccctggg caggggtctc caaatcccgg acgagccccc
     3961 aaatgaaaga cccccgtcgt gggtagtcaa tcactcagag gagaccctcc caaggaacag
     4021 cgagaccacg attcggatgc aaacagcaag aggctttatt gggaatacgg gtacccgggc
     4081 gacgcagtct atcggaggac tggcgcgccg agtgaggggt tgtgggctct tttattgagc
     4141 tctgcttata tagacctccc accgtacacg cctaccgccc atttgcgtca atggggcgga
     4201 gttgttacga cattttggaa agtcccgttg attttggtgc caaaacaaac tcccattgac
     4261 gtcaatgggg tggagacttg gaaatccccg tgagtcaaac cgctatccac gcccattgat
     4321 gtactgccaa aaccgcatca ccatggtaat agcgatgact aatacgtaga tgtactgcca
     4381 agtaggaaag tcccataagg tcatgtactg ggcataatgc caggcgggcc atttaccgtc
     4441 attgacgtca atagggggcg tacttggcat atgatacact tgatgtactg ccaagtgggc
     4501 agtttaccgt aaatactcca cccattgacg tcaatggaaa gtccctattg gcgttactat
     4561 gggaacatac gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc agccaggcgg
     4621 gccatttacc gtaagttatg taacgcggaa ctccatatat gggctatgaa ctaatgaccc
     4681 cgtaattgat tactattaat aactagtcaa taatcaatgt caacatggcg gtaatgttgg
     4741 acatgagcca atataaatgt acatattatg atatggatac aacgtatgca atggccaagc
     4801 taattcttga acacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     4861 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     4921 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     4981 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     5041 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     5101 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     5161 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     5221 taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg
     5281 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     5341 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     5401 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     5461 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     5521 cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa cgttgcgcaa
     5581 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     5641 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     5701 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     5761 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     5821 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     5881 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     5941 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     6001 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     6061 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     6121 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     6181 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     6241 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     6301 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     6361 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     6421 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     6481 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     6541 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     6601 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     6661 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     6721 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     6781 gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac
     6841 gcatctgtgc ggtatttcac accgcatatg gtgcactctc agtacaatct gctctgatgc
     6901 cgcatagtta agccagtaag ctttttgcaa aagcctaggc ctccaaaaaa gcctcctcac
     6961 tacttctgga atagctcaga ggccgaggcg gcctcggcct ctgcataaat aaaaaaaatt
     7021 agtcagccat ggggcggaga atgggcggaa ctgggcg
//
