LOCUS       sequence-207800-        6813 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shGAC (Target CCTCTGTTCTGTCAGAGTT), silence glutaminase isoform GAC,
            blasticidin selection..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6813)
  AUTHORS   Sandra Martha Gomes Dias
  TITLE     HuR (ELAVL1) silencing sensitizes triple-negative breast cancer to 
            glutaminase-targeted therapy
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6813)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pLKO.1-blast shGAC
FEATURES             Location/Qualifiers
     source          1..6813
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     240..260
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        242..260
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        288..514
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             515..695
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    742..867
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1360..1593
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1778..1822
                     /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"
     promoter        2120..2368
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2120..2140
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2291..2310
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2461..2578
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2627..3137
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3056..3074
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3149..3547
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             3619..3852
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3924..4045
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3961..3980)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4015..4034
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4085..4220
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4240..4259)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4241..4259)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4269..4286)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4269..4285)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4278..4300)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4427..4882
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4514..4533)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4724..4745
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4908..5012
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5013..5081,5082..5873)
                     /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(5231..5250)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6044..6632
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6533..6552
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6786..6803
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg
      241 caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgt agtcttatgc
      301 aatactcttg tagtcttgca acatggtaac gatgagttag caacatgcct tacaaggaga
      361 gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg ccttattagg
      421 aaggcaacag acgggtctga catggattgg acgaaccact gaattgccgc attgcagaga
      481 tattgtattt aagtgcctag ctcgatacat aaacgggtct ctctggttag accagatctg
      541 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
      601 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
      661 cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa
      721 gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg
      781 gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag
      841 aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg
      901 gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg
      961 gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc
     1021 tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga
     1081 tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac
     1141 accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag
     1201 caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt ggagaagtga
     1261 attatataaa tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa
     1321 gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt
     1381 cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg tacaggccag
     1441 acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca
     1501 acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa gaatcctggc
     1561 tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact
     1621 catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat
     1681 ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca caagcttaat
     1741 acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag aattattgga
     1801 attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc tgtggtatat
     1861 aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt ttgctgtact
     1921 ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga cccacctccc
     1981 aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag agagagacag
     2041 agacagatcc attcgattag tgaacggatc tcgacggtat cgatcacgag actagcctcg
     2101 agcggccgcc cccttcaccg agggcctatt tcccatgatt ccttcatatt tgcatatacg
     2161 atacaaggct gttagagaga taattggaat taatttgact gtaaacacaa agatattagt
     2221 acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt taaaattatg
     2281 ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt tcttggcttt
     2341 atatatcttg tggaaaggac gaaacaccgg cctctgttct gtcagagttc tcgagaactc
     2401 tgacagaaca gaggtttttg aattctcgac ctcgagacaa atggcagtat tcatccacaa
     2461 ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat
     2521 agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc aaaattttcg
     2581 ggtttattac agggacagca gagatccact ttggccgcgg ctcgaggggg ttggggttgc
     2641 gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg cgtggttccg
     2701 ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt cgcagcgtca
     2761 cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct ccgcccctaa
     2821 gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa gccgcacgtc
     2881 tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc cgaccgcgat
     2941 gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc cgggaagggg
     3001 cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc gcgcggtgtt
     3061 ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt gaccgaatca
     3121 ccgacctctc tccccagggg gatccaccat ggccaagcct ttgtctcaag aagaatccac
     3181 cctcattgaa agagcaacgg ctacaatcaa cagcatcccc atctctgaag actacagcgt
     3241 cgccagcgca gctctctcta gcgacggccg catcttcact ggtgtcaatg tatatcattt
     3301 tactggggga ccttgtgcag aactcgtggt gctgggcact gctgctgctg cggcagctgg
     3361 caacctgact tgtatcgtcg cgatcggaaa tgagaacagg ggcatcttga gcccctgcgg
     3421 acggtgccga caggtgcttc tcgatctgca tcctgggatc aaagccatag tgaaggacag
     3481 tgatggacag ccgacggcag ttgggattcg tgaattgctg ccctctggtt atgtgtggga
     3541 gggctaaggt acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
     3601 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa gatctgcttt
     3661 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     3721 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     3781 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     3841 aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga
     3901 aatgaatatc agagagtgag aggaacttgt ttattgcagc ttataatggt tacaaataaa
     3961 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     4021 tgtccaaact catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc
     4081 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     4141 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     4201 aggaggcttt tttggaggcc tagggacgta cccaattcgc cctatagtga gtcgtattac
     4261 gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     4321 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     4381 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc
     4441 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     4501 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     4561 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     4621 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     4681 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     4741 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     4801 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     4861 aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     4921 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     4981 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     5041 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     5101 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     5161 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     5221 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     5281 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     5341 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     5401 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     5461 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     5521 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     5581 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     5641 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     5701 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     5761 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     5821 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     5881 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     5941 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     6001 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     6061 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     6121 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     6181 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     6241 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     6301 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     6361 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     6421 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     6481 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     6541 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     6601 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     6661 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     6721 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     6781 agcgcagcga gtcagtgagc gaggaagcgg aag
//
