LOCUS       sequence-251619-        7983 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A TCR Variable Region cloning expression vector using Mouse Constant
            Regions to reduce mis-pairing with endogenous TCR expressing NGFR 
            ref Biotechniques. 2015 Mar 1;58(3):135-9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7983)
  AUTHORS   Coren LV, Jain S, Trivett MT, Ohlen C, Ott DE
  TITLE     Production of retroviral constructs for effective transfer and 
            expression of T-cell receptor genes using Golden Gate cloning.
  JOURNAL   Biotechniques. 2015 Mar 1;58(3):135-9. doi: 10.2144/000114265. 
            eCollection 2015.
  PUBMED    25757546
REFERENCE   2  (bases 1 to 7983)
  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: "10.2144/000114265";
            journalName: "Biotechniques"; date: "2015-03-1- 1"; volume: "58"; 
            issue: "3"; pages: "135-9"
COMMENT     Sequence Label: MSGV Mouse Acceptor PGK NGFR
FEATURES             Location/Qualifiers
     source          1..7983
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..517
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     primer_bind     1357..1379
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1395..1411
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     1749..1765
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     promoter        2892..3391
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2927..2950)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3324..3343
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     misc_feature    4272..4324
                     /label=MCS
                     /note="multiple cloning site"
     LTR             4368..4882
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5054..5070
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5078..5084,5085..5102,5103..5108))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5123..5144
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5261..5278)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5432..6020)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5512..5531)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6191..6982,6983..7051))
                     /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     6814..6833
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7052..7156)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7224..7242
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7280..7302)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7402..7421
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7615..7637
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7765..7784
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag
      121 cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
      181 gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt
      241 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
      301 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
      361 ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc gtattcccaa
      421 taaagcctct tgctgtttgc atccgaatcg tggactcgct gatccttggg agggtctcct
      481 cagattgatt gactgcccac ctcgggggtc tttcatttgg aggttccacc gagatttgga
      541 gacccctgcc cagggaccac cgaccccccc gccgggaggt aagctggcca gcggtcgttt
      601 cgtgtctgtc tctgtctttg tgcgtgtttg tgccggcatc taatgtttgc gcctgcgtct
      661 gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg acgagttcgg
      721 aacacccggc cgcaaccctg ggagacgtcc cagggacttc gggggccgtt tttgtggccc
      781 gacctgagtc ctaaaatccc gatcgtttag gactctttgg tgcacccccc ttagaggagg
      841 gatatgtggt tctggtagga gacgagaacc taaaacagtt cccgcctccg tctgaatttt
      901 tgctttcggt ttgggaccga agccgcgccg cgcgtcttgt ctgctgcagc atcgttctgt
      961 gttgtctctg tctgactgtg tttctgtatt tgtctgaaaa tatgggcccg ggctagcctg
     1021 ttaccactcc cttaagtttg accttaggtc actggaaaga tgtcgagcgg atcgctcaca
     1081 accagtcggt agatgtcaag aagagacgtt gggttacctt ctgctctgca gaatggccaa
     1141 cctttaacgt cggatggccg cgagacggca cctttaaccg agacctcatc acccaggtta
     1201 agatcaaggt cttttcacct ggcccgcatg gacacccaga ccaggtcccc tacatcgtga
     1261 cctgggaagc cttggctttt gacccccctc cctgggtcaa gccctttgta caccctaagc
     1321 ctccgcctcc tcttcctcca tccgccccgt ctctccccct tgaacctcct cgttcgaccc
     1381 cgcctcgatc ctccctttat ccagccctca ctccttctct aggcgccccc atatggccat
     1441 atgagatctt atatggggca cccccgcccc ttgtaaactt ccctgaccct gacatgacaa
     1501 gagttactaa cagcccctct ctccaagctc acttacaggc tctctactta gtccagcacg
     1561 aagtctggag acctctggcg gcagcctacc aagaacaact ggaccgaccg gtggtacctc
     1621 acccttaccg agtcggcgac acagtgtggg tccgccgaca ccagactaag aacctagaac
     1681 ctcgctggaa aggaccttac acagtcctgc tgaccacccc caccgccctc aaagtagacg
     1741 gcatcgcagc ttggatacac gccgcccacg tgaaggctgc cgaccccggg ggtggaccat
     1801 cctctagacc gcaacgttaa ttaagccacc aagtcttctt tttttgaaga cttcagaacc
     1861 ccgagcccgc cgtgtaccag ctgaaggacc ccagaagcca ggacagcacc ctgtgtctgt
     1921 tcaccgactt cgacagccag atcaacgtgc ccaagaccat ggaaagcggc accttcatca
     1981 ccgataagtg cgtgctggac atgaaggcca tggacagcaa gagcaacggc gccattgcct
     2041 ggtccaacca gaccagcttc acatgccagg acatcttcaa agagacaaac gccacctacc
     2101 cttccagcga cgtgccctgt gacgccaccc tgaccgagaa gtccttcgag acagacatga
     2161 atctgaattt ccagaacctg agcgtgatgg gcctgcggat cctgctgctg aaagtggccg
     2221 gcttcaacct gctgatgacc ctgcggctgt ggtccagccg ggccaagaga tctggcagcg
     2281 gcgccaccaa tttcagcctg ctgaaacagg ccggggacgt ggaagagaac cctaagtctt
     2341 ctttttgaag acttgatctg aggaacgtga ccccccccaa ggtgtccctg ttcgagccca
     2401 gcaaggccga gatcgccaac aagcagaaag ccaccctcgt gtgcctggcc agaggcttct
     2461 tccccgacca cgtggaactg tcctggtggg tcaacggcaa agaggtgcac agcggagtct
     2521 gcaccgaccc ccaggcttac aaagagagca actacagcta ctgcctgtcc agcagactgc
     2581 gggtgtccgc taccttctgg cacaaccccc ggaaccactt cagatgccag gtgcagttcc
     2641 acggcctgag cgaagaggac aagtggcccg agggcagccc caagcccgtg acccagaata
     2701 tcagcgccga ggcctggggc agagccgatt gtggcatcac cagcgccagc taccaccagg
     2761 gggtgctgag cgccaccatc ctgtacgaga tcctgctggg caaggccacc ctgtacgccg
     2821 tgctggtgtc cggactggtg ctgatggcca tggtcaagaa gaagaacagc tagcggccgc
     2881 tacaaattac cgggtagggg aggcgctttt cccaaggcag tctggagcat gcgctttagc
     2941 agccccgctg ggcacttggc gctacacaag tggcctctgg cctcgcacac attccacatc
     3001 caccggtagg cgccaaccgg ctccgttctt tggtggcccc ttcgcgccac cttctactcc
     3061 tcccctagtc aggaagttcc cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat
     3121 ggaagtagca cgtctcacta gtctcgtgca gatggacagc accgctgagc aatggaagcg
     3181 ggtaggcctt tggggcagcg gccaatagca gctttgctcc ttcgctttct gggctcagag
     3241 gctgggaagg ggtgggtccg ggggcgggct caggggcggg ctcaggggcg gggcgggcgc
     3301 ccgaaggtcc tccggaggcc cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct
     3361 gttctcctct tcctcatctc cgggcctttc gacctgcagg cggccgcacc atgggggcag
     3421 gtgccactgg ccgcgccatg gacgggccgc gcctgctgct gttgctgctt ctgggggtgt
     3481 cccttggagg tgccaaggag gcatgcccca caggcctgta cacacacagc ggtgagtgct
     3541 gcaaagcctg caacctgggc gagggtgtgg cccagccttg tggagccaac cagaccgtgt
     3601 gtgagccctg cctggacagc gtgacgttct ccgacgtggt gagcgcgacc gagccgtgca
     3661 agccgtgcac cgagtgcgtg gggctccaga gcatgtcggc gccatgcgtg gaggccgacg
     3721 acgccgtgtg ccgctgcgcc tacggctact accaggatga gacgactggg cgctgcgagg
     3781 cgtgccgcgt gtgcgaggcg ggctcgggcc tcgtgttctc ctgccaggac aagcagaaca
     3841 ccgtgtgcga ggagtgcccc gacggcacgt attccgacga ggccaaccac gtggacccgt
     3901 gcctgccctg caccgtgtgc gaggacaccg agcgccagct ccgcgagtgc acacgctggg
     3961 ccgacgccga gtgcgaggag atccctggcc gttggattac acggtccaca cccccagagg
     4021 gctcggacag cacagccccc agcacccagg agcctgaggc acctccagaa caagacctca
     4081 tagccagcac ggtggcaggt gtggtgacca cagtgatggg cagctcccag cccgtggtga
     4141 cccgaggcac caccgacaac ctcatccctg tctattgctc catcctggct gctgtggttg
     4201 tgggtcttgt ggcctacata gccttcaaga ggtggaacag ctgagcggcc gctacgtaga
     4261 tcgacggatc cctcgagctc aagcttcgaa ttctgcagtc gacggtaccg cgggcccggg
     4321 atccgataaa ataaaagatt ttatttagtc tccagaaaaa ggggggaatg aaagacccca
     4381 cctgtaggtt tggcaagcta gcttaagtaa cgccattttg caaggcatgg aaaatacata
     4441 actgagaata gagaagttca gatcaaggtt aggaacagag agacagcaga atatgggcca
     4501 aacaggatat ctgtggtaag cagttcctgc cccggctcag ggccaagaac agatggtccc
     4561 cagatgcggt cccgccctca gcagtttcta gagaaccatc agatgtttcc agggtgcccc
     4621 aaggacctga aatgaccctg tgccttattt gaactaacca atcagttcgc ttctcgcttc
     4681 tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc cacaacccct cactcggcgc
     4741 gccagtcctc cgatagactg cgtcgcccgg gtacccgtgt atccaataaa ccctcttgca
     4801 gttgcatccg acttgtggtc tcgctgttcc ttgggagggt ctcctctgag tgattgacta
     4861 cccgtcagcg ggggtctttc atgggtaaca gtttcttgaa gttggagaac aacattctga
     4921 gggtaggagt cgaatattaa gtaatcctga ctcaattagc cactgttttg aatccacata
     4981 ctccaatact cctgaaatcc atcgatggag ttcattatgg acagcgcaga aagagctggg
     5041 gagaattgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     5101 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     5161 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     5221 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     5281 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     5341 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     5401 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     5461 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     5521 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     5581 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     5641 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     5701 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     5761 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     5821 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     5881 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5941 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     6001 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     6061 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     6121 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     6181 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     6241 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     6301 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     6361 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     6421 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     6481 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     6541 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6601 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6661 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6721 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6781 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6841 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6901 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6961 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     7021 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     7081 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     7141 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta
     7201 ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt
     7261 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc
     7321 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt
     7381 gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc
     7441 ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc cattcgccat
     7501 tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc
     7561 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     7621 cacgacgttg taaaacgacg gcgcaaggaa tggtgcatgc aaggagatgg cgcccaacag
     7681 tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa
     7741 gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc
     7801 tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggcgattag tccaatttgt
     7861 taaagacagg atatcagtgg tccaggctct agttttgact caacaatatc accagctgaa
     7921 gcctatagag tacgagccat agataaaata aaagatttta tttagtctcc agaaaaaggg
     7981 ggg
//
