LOCUS       sequence-379963-        4331 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses KRAS in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4331)
  AUTHORS   Inoue M, Baba T, Takahashi F, Terao M, Yanai S, Shima Y, Saito D, 
            Sugihara K, Miura T, Takada S, Suyama M, Ohkawa Y, Morohashi KI
  TITLE     Tmsb10 triggers fetal Leydig differentiation by suppressing the 
            RAS/ERK pathway.
  JOURNAL   Commun Biol. 2022 Sep 15;5(1):974. doi: 10.1038/s42003-022-03941-5.
  PUBMED    36109592
REFERENCE   2  (bases 1 to 4331)
  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: "Commun 
            Biol."; date: "2022-09-15"; volume: "5(1):974. doi"; pages: " 
            10.1038/s42003-022-03941-5"
COMMENT     Sequence Label: pCMV-HA-KRAS
FEATURES             Location/Qualifiers
     source          1..4331
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        40..343
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        344..547
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     498..518
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     544..568
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          685..781
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             845..871
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    1483..1617
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1507..1526)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1561..1580
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1728..1744)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1728..1744)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1741..1763)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1752..1768
                     /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(1776..1782,1783..1800,1801..1806))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1821..1842
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1959..1976)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2130..2718)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2210..2229)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2889..3680,3681..3749))
                     /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     3512..3531
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3750..3854)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3922..3940
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3978..4000)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4100..4119
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(4313..4331,1..4)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(4327..4331,1..13)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     join(4328..4331,1..13)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 aacgacggcc agtgagttcg agcttgcatg cctgcaggtc gttacataac ttacggtaaa
       61 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      121 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      181 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      241 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      301 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
      361 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
      421 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
      481 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
      541 cagagctcgt ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct
      601 ccatagaaga caccgggacc gatccagcct ccggactcta gaggatccgg tactagagga
      661 actgaaaaac cagaaagtta actggtaagt ttagtctttt tgtcttttat ttcaggtccc
      721 ggatccggtg gtggtgcaaa tcaaagaact gctcctcagt ggatgttgcc tttacttcta
      781 ggcctgtacg gaagtgttac ttctgctcta aaagctgcgg aattgtaccc gcgggcccac
      841 catgtaccca tacgatgttc cagattacgc tcttatggcc atggaggccc gaattcggtc
      901 gaccatgact gagtataaac ttgtggtggt tggagctggt ggcgtaggca agagcgcctt
      961 gacgatacag ctaattcaga atcactttgt ggatgagtac gaccctacga tagaggactc
     1021 ctacaggaaa caagtagtaa ttgatggaga aacctgtctc ttggatattc tcgacacagc
     1081 aggtcaagag gagtacagtg caatgaggga ccagtacatg agaactgggg agggctttct
     1141 ttgtgtattt gccataaata atactaaatc atttgaagat attcaccatt atagagaaca
     1201 aattaaaaga gtaaaggact ctgaagatgt gcctatggtc ctggtaggga ataagtgtga
     1261 tttgccttct agaacagtag acacgaaaca ggctcaggag ttagcaagga gttacgggat
     1321 tccgttcatt gagacctcag caaagacaag acagggtgtt gacgatgcct tctatacatt
     1381 agtccgagaa attcgaaaac ataaagaaaa gatgagcaaa gatgggaaga agaagaagaa
     1441 gaagtcaagg acaaggtgta cagttatgtg agcggccgcg gggatccaga catgataaga
     1501 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
     1561 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttaac
     1621 aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttcg
     1681 gatcctctag agtcgatctg caggcatgct agcttggcgt aatcatggtc atagctgttt
     1741 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     1801 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     1861 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     1921 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     1981 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     2041 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     2101 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     2161 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     2221 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     2281 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     2341 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     2401 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     2461 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     2521 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     2581 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     2641 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     2701 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     2761 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     2821 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     2881 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     2941 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     3001 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     3061 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     3121 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     3181 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     3241 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     3301 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     3361 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     3421 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     3481 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     3541 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     3601 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     3661 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     3721 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     3781 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     3841 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     3901 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc
     3961 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     4021 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     4081 cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg
     4141 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgcca ttcgccattc
     4201 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
     4261 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
     4321 cgacgttgta a
//
