LOCUS       sequence-422379-        8001 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Retroviral golden gate cloning vector for expression of functional 
            Human T-Cell Receptor genes with NGFR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8001)
  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 8001)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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 Hu Acceptor PGK-NGFR
FEATURES             Location/Qualifiers
     source          1..8001
                     /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        2910..3409
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2945..2968)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3342..3361
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     misc_feature    4290..4342
                     /label=MCS
                     /note="multiple cloning site"
     LTR             4386..4900
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5072..5088
                     /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(5096..5102,5103..5120,5121..5126))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5141..5162
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5279..5296)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5450..6038)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5530..5549)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6209..7000,7001..7069))
                     /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     6832..6851
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7070..7174)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7242..7260
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7298..7320)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7420..7439
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7633..7655
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7783..7802
                     /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 ctgatcctgc cgtgtaccag ctgcgggaca gcaagagcag cgacaagagc gtgtgcctgt
     1921 tcaccgactt cgacagccag accaacgtgt cccagagcaa ggacagcgac gtgtacatca
     1981 ccgacaagac cgtgctggac atgcggagca tggacttcaa gagcaacagc gccgtggcct
     2041 ggtccaacaa gagcgatttc gcctgcgcca acgccttcaa caacagcatt atccccgagg
     2101 acacattctt cccaagcccc gagagcagct gcgacgtgaa gctggtggaa aagagcttcg
     2161 agacagacac caacctgaac ttccagaacc tgagcgtgat cggcttccgg atcctgctgc
     2221 tgaaggtggc cggcttcaac ctgctgatga ccctgagact gtggtccagc cgggccaaga
     2281 gatctggcag cggcgccacc aatttcagcc tgctgaaaca ggccggcgac gtggaagaga
     2341 accctaagtc ttctttttga agacttgatc tgaacaaggt gttcccccca gaggtggccg
     2401 tgttcgagcc ttctgaggcc gagatcagcc acacccagaa agccaccctc gtgtgcctgg
     2461 ccaccggctt tttccccgac cacgtggaac tgtcttggtg ggtcaacggc aaagaggtgc
     2521 acagcggcgt gtccaccgat ccccagcctc tgaaagaaca gcccgccctg aacgacagcc
     2581 ggtactgcct gagcagcaga ctgagagtgt ccgccacctt ctggcagaac ccccggaacc
     2641 acttcagatg ccaggtgcag ttctacggcc tgagcgagaa cgacgagtgg acccaggaca
     2701 gagccaagcc cgtgacccag atcgtgtctg ccgaagcctg gggcagagcc gattgcggct
     2761 ttaccagcgt gtcctatcag cagggcgtgc tgagcgccac catcctgtac gagatcctgc
     2821 tgggcaaggc caccctgtac gccgtgctgg tgtctgccct ggtgctgatg gccatggtca
     2881 agcggaagga cttctgaatt taaattaccg ggtaggggag gcgcttttcc caaggcagtc
     2941 tggagcatgc gctttagcag ccccgctggg cacttggcgc tacacaagtg gcctctggcc
     3001 tcgcacacat tccacatcca ccggtaggcg ccaaccggct ccgttctttg gtggcccctt
     3061 cgcgccacct tctactcctc ccctagtcag gaagttcccc cccgccccgc agctcgcgtc
     3121 gtgcaggacg tgacaaatgg aagtagcacg tctcactagt ctcgtgcaga tggacagcac
     3181 cgctgagcaa tggaagcggg taggcctttg gggcagcggc caatagcagc tttgctcctt
     3241 cgctttctgg gctcagaggc tgggaagggg tgggtccggg ggcgggctca ggggcgggct
     3301 caggggcggg gcgggcgccc gaaggtcctc cggaggcccg gcattctgca cgcttcaaaa
     3361 gcgcacgtct gccgcgctgt tctcctcttc ctcatctccg ggcctttcga cctgcaggcg
     3421 gccgcaccat gggggcaggt gccactggcc gcgccatgga cgggccgcgc ctgctgctgt
     3481 tgctgcttct gggggtgtcc cttggaggtg ccaaggaggc atgccccaca ggcctgtaca
     3541 cacacagcgg tgagtgctgc aaagcctgca acctgggcga gggtgtggcc cagccttgtg
     3601 gagccaacca gaccgtgtgt gagccctgcc tggacagcgt gacgttctcc gacgtggtga
     3661 gcgcgaccga gccgtgcaag ccgtgcaccg agtgcgtggg gctccagagc atgtcggcgc
     3721 catgcgtgga ggccgacgac gccgtgtgcc gctgcgccta cggctactac caggatgaga
     3781 cgactgggcg ctgcgaggcg tgccgcgtgt gcgaggcggg ctcgggcctc gtgttctcct
     3841 gccaggacaa gcagaacacc gtgtgcgagg agtgccccga cggcacgtat tccgacgagg
     3901 ccaaccacgt ggacccgtgc ctgccctgca ccgtgtgcga ggacaccgag cgccagctcc
     3961 gcgagtgcac acgctgggcc gacgccgagt gcgaggagat ccctggccgt tggattacac
     4021 ggtccacacc cccagagggc tcggacagca cagcccccag cacccaggag cctgaggcac
     4081 ctccagaaca agacctcata gccagcacgg tggcaggtgt ggtgaccaca gtgatgggca
     4141 gctcccagcc cgtggtgacc cgaggcacca ccgacaacct catccctgtc tattgctcca
     4201 tcctggctgc tgtggttgtg ggtcttgtgg cctacatagc cttcaagagg tggaacagct
     4261 gagcggccgc tacgtagatc gacggatccc tcgagctcaa gcttcgaatt ctgcagtcga
     4321 cggtaccgcg ggcccgggat ccgataaaat aaaagatttt atttagtctc cagaaaaagg
     4381 ggggaatgaa agaccccacc tgtaggtttg gcaagctagc ttaagtaacg ccattttgca
     4441 aggcatggaa aatacataac tgagaataga gaagttcaga tcaaggttag gaacagagag
     4501 acagcagaat atgggccaaa caggatatct gtggtaagca gttcctgccc cggctcaggg
     4561 ccaagaacag atggtcccca gatgcggtcc cgccctcagc agtttctaga gaaccatcag
     4621 atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg ccttatttga actaaccaat
     4681 cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc gagctcaata aaagagccca
     4741 caacccctca ctcggcgcgc cagtcctccg atagactgcg tcgcccgggt acccgtgtat
     4801 ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc gctgttcctt gggagggtct
     4861 cctctgagtg attgactacc cgtcagcggg ggtctttcat gggtaacagt ttcttgaagt
     4921 tggagaacaa cattctgagg gtaggagtcg aatattaagt aatcctgact caattagcca
     4981 ctgttttgaa tccacatact ccaatactcc tgaaatccat cgatggagtt cattatggac
     5041 agcgcagaaa gagctgggga gaattgtgaa attgttatcc gctcacaatt ccacacaaca
     5101 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     5161 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     5221 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     5281 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5341 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     5401 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5461 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5521 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5581 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5641 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5701 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5761 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5821 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5881 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5941 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     6001 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     6061 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     6121 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6181 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6241 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6301 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6361 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6421 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6481 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6541 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6601 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6661 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6721 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6781 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6841 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6901 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6961 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     7021 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     7081 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     7141 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7201 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     7261 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     7321 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     7381 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac
     7441 tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     7501 tcaggcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct
     7561 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
     7621 cgccagggtt ttcccagtca cgacgttgta aaacgacggc gcaaggaatg gtgcatgcaa
     7681 ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca
     7741 agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata
     7801 ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag
     7861 gcgattagtc caatttgtta aagacaggat atcagtggtc caggctctag ttttgactca
     7921 acaatatcac cagctgaagc ctatagagta cgagccatag ataaaataaa agattttatt
     7981 tagtctccag aaaaaggggg g
//
