LOCUS       Exported                7057 bp ds-DNA     circular SYN 16-NOV-2021
DEFINITION  knockout mouse CD4.
ACCESSION   .
VERSION     .
KEYWORDS    pSIN-CD4
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 Nov 16, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7057
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     62..81
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(162..179)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(333..921)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(413..432)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1092..1952)
                     /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     1715..1734
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1953..2057)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2125..2143
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        2243..2622
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2623..2826
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2777..2797
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             2827..3002
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    3065..3264
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             3465..3880
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     3810..3832
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     3848..3864
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        3927..4167
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     3927..3947
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     4098..4117
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        4197..4272
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        4379..4590
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             4610..5095
                     /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"
     CDS             5105..5161
                     /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             5162..5956
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5216..5235)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5826..5845
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     LTR             6077..6502
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(6605..6627)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6763..6783)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6768..40
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6948..26
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7010..7029
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagcttactg gcttaactat
       61 gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga
      121 tgcgtaagga gaaaataccg catcaggcgc tcttccgctt cctcgctcac tgactcgctg
      181 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
      241 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
      301 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
      361 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
      421 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
      481 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
      541 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
      601 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
      661 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
      721 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
      781 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
      841 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
      901 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
      961 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     1021 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     1081 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     1141 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     1201 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     1261 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     1321 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     1381 agtttgcgca acgttgttgc cattgctgca ggcatcgtgg tgtcacgctc gtcgtttggt
     1441 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     1501 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     1561 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     1621 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     1681 cgaccgagtt gctcttgccc ggcgtcaaca cgggataata ccgcgccaca tagcagaact
     1741 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     1801 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     1861 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     1921 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     1981 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     2041 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt
     2101 attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg tgttcaagaa
     2161 ttagcttggc cattgcatac gttgtatcca tatcataata tgtacattta tattggctca
     2221 tgtccaacat taccgccatg ttgacattga ttattgacta gttattaata gtaatcaatt
     2281 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     2341 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     2401 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     2461 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     2521 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     2581 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     2641 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     2701 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     2761 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     2821 agagctcaat aaaagagccc acaacccctc actcggcgcg ccagtcctcc gatagactgc
     2881 gtcgcccggg tacccgtatt cccaataaag cctcttgctg tttgcatccg aatcgtggtc
     2941 tcgctgttcc ttgggagggt ctcctctgag tgattgacta cccacgacgg gggtctttca
     3001 tttgggggct cgtccgggat ttggagaccc ctgcccaggg accaccgacc caccaccggg
     3061 aggtaagctg gccagcaact tatctgtgtc tgtccgattg tctagtgtct atgtttgatg
     3121 ttatgcgcct gcgtctgtac tagttagcta actagctctg tatctggcgg acccgtggtg
     3181 gaactgacga gttctgaaca cccggccgca accctgggag acgtcccagg gactttgggg
     3241 gccgtttttg tggcccgacc tgaggaaggg agtcgatgtg gaatccgacc ccgtcaggat
     3301 atgtggttct ggtaggagac gagaacctaa aacagttccc gcctccgtct gaatttttgc
     3361 tttcggtttg gaaccgaagc cgcgcgtctt gtctgctgca gcgctgcagc atcgttctgt
     3421 gttgtctctg tctgactgtg tttctgtatt tgtctgaaaa ttagggccag actgttacca
     3481 ctcccttaag tttgacctta ggtcactgga aagatgtcga gcggatcgct cacaaccagt
     3541 cggtagatgt caagaagaga cgttggttac cttctgctct gcagaatggc caacctttaa
     3601 cgtcggatgg ccgcgagacg gcacctttaa ccgagacctc atcacccagg ttaagatcaa
     3661 ggtcttttca cctggcccgc atggacaccc agaccaggtc ccctacatcg tgacctggga
     3721 agccttggct tttgaccccc ctccctgggt caagcccttt gtacacccta agcctccgcc
     3781 tcctcttcct ccatccgccc cgtctctccc ccttgaacct cctcgttcga ccccgcctcg
     3841 atcctccctt tatccagccc tcactccttc tctaggcgcc ggaattgaag atctttaatt
     3901 aacgatctat cgattctaga tctagagagg gcctatttcc catgattcct tcatatttgc
     3961 atatacgata caaggctgtt agagagataa ttagaattaa tttgactgta aacacaaaga
     4021 tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt gcagttttaa
     4081 aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct
     4141 tggctttata tatcttgtgg aaaggacgaa acaccgcctg gaagttctct gaccaggttt
     4201 tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga aaaagtggca
     4261 ccgagtcggt gcttttttga atttagatct tgagacaagg tgccgggccg ggaattcggc
     4321 gaattgggcc cgacgtcgca tgctcccggc ccctcgagtg gctccggtgc ccgtcagtgg
     4381 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc
     4441 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     4501 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     4561 tttcgcaacg ggtttgccgc cagaacacag gtgtcgtgac gcggccacca tgaacccagc
     4621 catcagcgtc gctctcctgc tctcagtctt gcaggtgtcc cgagggcaga aggtgaccag
     4681 cctgacagcc tgcctggtga accaaaacct tcgcctggac tgccgccatg agaataacac
     4741 caaggataac tccatccagc atgagttcag cctgacccga gagaagagga agcacgtgct
     4801 ctcaggcacc ctcgggatac ccgagcacac gtaccgctcc cgcgtcaccc tctccaacca
     4861 gccctatatc aaggtcctta ccctagccaa cttcaccacc aaggatgagg gcgactactt
     4921 ttgtgagctt cgagtctcgg gcgcgaatcc catgagctcc aataaaagta tcagtgtgta
     4981 tagagacaaa ctggtcaagt gtggcggcat aagcctgctg gttcagaaca catcctggat
     5041 gctgctgctg ctgctttccc tctccctcct ccaagccctg gacttcattt ctctgggcag
     5101 cggcgccacc aacttcagcc tgctgaagca ggccggcgac gtggaggaga accccggccc
     5161 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     5221 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     5281 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     5341 gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     5401 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     5461 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     5521 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     5581 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     5641 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga
     5701 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     5761 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     5821 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     5881 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     5941 tgacgagttc ttctgattag atatcatcag tggtccaggc tctagttttg actcaacaat
     6001 atcaccagct gaagcctata gagtacgagc catagataaa ataaaagatt ttatttagtc
     6061 tccagaaaaa ggggggaatg aaagacccca cctgtaggtt tggcaagcta gcttaagtaa
     6121 cgccattttg caaggcatgg aaaaatacat aactgagaat agagaagttc agatcaaggt
     6181 caggaacaga tggaacaggg tcgaccctag agaaccatca gatgtttcca gggtgcccca
     6241 aggacctgaa atgaccctgt gccttatttg aactaaccaa tcagttcgct tctcgcttct
     6301 gttcgcgcgc ttctgctccc cgagctcaat aaaagagccc acaacccctc actcggggcg
     6361 ccagtcctcc gattgactga gtcgcccggg tacccgtgta tccaataaac cctcttgcag
     6421 ttgcatccga cttgtggtct cgctgttcct tgggagggtc tcctctgagt gattgactac
     6481 ccgtcagcgg gggtctttca tttgggggct cgtccgggat cgggagaccc ctgcccaggg
     6541 accaccgacc caccaccggg aggtaagctg gctgcctcgc gcgtttcggt gatgacggtg
     6601 aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg
     6661 ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca
     6721 tgacccagtc acgtagcgat agcggagtgt agatccggct gtggaatgtg tgtcagttag
     6781 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     6841 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     6901 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     6961 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     7021 aggccgaggc cgcctcggcc tctgagctat tccagaa
//
