LOCUS       sequence-192205-        7084 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Knockdown of MCT4 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  AUTHORS   Lim KS, Lim KJ, Price AC, Orr BA, Eberhart CG, Bar EE
  TITLE     Inhibition of monocarboxylate transporter-4 depletes stem-like 
            glioblastoma cells and inhibits HIF transcriptional response in a 
            lactate-independent manner.
  JOURNAL   Oncogene. 2014 Aug 28;33(35):4433-41. doi: 10.1038/onc.2013.390. 
            Epub 2013 Sep 30.
  PUBMED    24077291
REFERENCE   2  (bases 1 to 7084)
  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: 
            "10.1038/onc.2013.390"; journalName: "Oncogene"; date: "2014-08-28- 
            28"; volume: "33"; issue: "35"; pages: "4433-41"
COMMENT     Sequence Label: plko.1/shMCT4 #474
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(155..174)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(156..174)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(184..201)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(184..200)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(193..215)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      342..797
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(429..448)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     639..660
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        823..927
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(928..996,997..1788)
                     /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(1146..1165)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1959..2547
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2448..2467
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2701..2718
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2835..2856
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2871..2876,2877..2894,2895..2901)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2909..2925
                     /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     2914..2936
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2933..2949
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2933..2949
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2968..2988
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2970..2988
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3016..3242
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             3243..3423
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    3470..3595
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    4088..4321
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             4506..4550
                     /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        4848..5096
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     4848..4868
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     5019..5038
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    5193..5310
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5359..5869
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     5788..5806
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             5891..6490
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5891..5910)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6387..6407
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             6618..6851
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6923..7044
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6960..6979)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7014..7033
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      join(7084,1..135)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
       61 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
      121 gcttttttgg aggcctaggg acgtacccaa ttcgccctat agtgagtcgt attacgcgcg
      181 ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
      241 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
      301 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc
      361 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
      421 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
      481 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
      541 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
      601 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
      661 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
      721 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
      781 attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
      841 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
      901 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
      961 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     1021 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     1081 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     1141 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     1201 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     1261 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     1321 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     1381 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     1441 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     1501 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     1561 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     1621 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     1681 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     1741 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     1801 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     1861 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     1921 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     1981 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2041 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2101 actgttcttc tagtgtatcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2161 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     2221 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     2281 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     2341 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     2401 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     2461 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     2521 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     2581 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     2641 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     2701 agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg
     2761 cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt
     2821 gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc tttacacttt
     2881 atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac
     2941 agctatgacc atgattacgc caagcgcgca attaaccctc actaaaggga acaaaagctg
     3001 gagctgcaag cttaatgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga
     3061 tgagttagca acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag
     3121 taaggtggta cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac
     3181 gaaccactga attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa
     3241 acgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc
     3301 cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt
     3361 tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta
     3421 gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg
     3481 caggactcgg cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac
     3541 gccaaaaatt ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat
     3601 taagcggggg agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa
     3661 aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa
     3721 tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc
     3781 ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg
     3841 tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga
     3901 gcaaaacaaa agtaagacca ccgcacagca agcggccgct gatcttcaga cctggaggag
     3961 gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac
     4021 cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag
     4081 tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag
     4141 cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga
     4201 acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca
     4261 tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc
     4321 tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta
     4381 gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca
     4441 gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc
     4501 aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt
     4561 ttaacataac aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg
     4621 taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt
     4681 caccattatc gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa
     4741 tagaagaaga aggtggagag agagacagag acagatccat tcgattagtg aacggatctc
     4801 gacggtatcg atcacgagac tagcctcgag cggccgcccc cttcaccgag ggcctatttc
     4861 ccatgattcc ttcatatttg catatacgat acaaggctgt tagagagata attggaatta
     4921 atttgactgt aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc
     4981 ttgggtagtt tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa
     5041 cttgaaagta tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccggcg
     5101 tctacatgta cgtgttcatc tcgagatgaa cacgtacatg tagacgtttt tgaattctcg
     5161 acctcgagac aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg
     5221 tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta
     5281 caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca
     5341 ctttggccgc ggctcgaggg ggttggggtt gcgccttttc caaggcagcc ctgggtttgc
     5401 gcagggacgc ggctgctctg ggcgtggttc cgggaaacgc agcggcgccg accctgggtc
     5461 tcgcacattc ttcacgtccg ttcgcagcgt cacccggatc ttcgccgcta cccttgtggg
     5521 ccccccggcg acgcttcctg ctccgcccct aagtcgggaa ggttccttgc ggttcgcggc
     5581 gtgccggacg tgacaaacgg aagccgcacg tctcactagt accctcgcag acggacagcg
     5641 ccagggagca atggcagcgc gccgaccgcg atgggctgtg gccaatagcg gctgctcagc
     5701 agggcgcgcc gagagcagcg gccgggaagg ggcggtgcgg gaggcggggt gtggggcggt
     5761 agtgtgggcc ctgttcctgc ccgcgcggtg ttccgcattc tgcaagcctc cggagcgcac
     5821 gtcggcagtc ggctccctcg ttgaccgaat caccgacctc tctccccagg gggatccacc
     5881 ggagcttacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc
     5941 cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt
     6001 cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt
     6061 cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac
     6121 cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga
     6181 gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg
     6241 gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa
     6301 gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc
     6361 cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac
     6421 cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc
     6481 cggtgcctga cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg cacgacccca
     6541 tgcatcggta cctttaagac caatgactta caaggcagct gtagatctta gccacttttt
     6601 aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag atctgctttt
     6661 tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     6721 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     6781 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     6841 aatctctagc agtagtagtt catgtcatct tattattcag tatttataac ttgcaaagaa
     6901 atgaatatca gagagtgaga ggaacttgtt tattgcagct tataatggtt acaaataaag
     6961 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     7021 gtccaaactc atcaatgtat cttatcatgt ctggctctag ctatcccgcc cctaactccg
     7081 ccca
//
