LOCUS       sequence-502963-        6853 bp    DNA     circular SYN 13-MAR-2026
DEFINITION  Bicistronic expression of Histone2B-mCherry fusion and Hygro 
            resistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6853)
  TITLE     Piel Lab Plasmids
REFERENCE   2  (bases 1 to 6853)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Mar 13, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: Histone2B mCherry Hygro
FEATURES             Location/Qualifiers
     source          1..6853
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             696..1073
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFEHIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             1080..1784
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELY"
     primer_bind     complement(1222..1240)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1490..1509
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     intron          1889..2118
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2101..2120
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(2201..2775,2776..2778,2779..2787)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2368..2385)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2595..2614
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2813..3832
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     CDS             2813..3832
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     polyA_signal    4240..4361
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4277..4296)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4462..4478)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4462..4478)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4475..4497)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4486..4502
                     /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(4510..4516,4517..4534,4535..4540))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4555..4576
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4693..4710)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4864..5452)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4944..4963)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5623..6414,6415..6483))
                     /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     6246..6265
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6484..6588)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6663..6682)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccgagc tcggatctgt acaggcctta agaccatgcc agagccagcg aagtctgctc
      721 ccgccccgaa aaagggctcc aagaaggcgg tgactaaggc gcagaagaaa ggcggcaaga
      781 agcgcaagcg cagccgcaag gagagctatt ccatctatgt gtacaaggtt ctgaagcagg
      841 tccaccctga caccggcatt tcgtccaagg ccatgggcat catgaattcg tttgtgaacg
      901 acattttcga gcacatcgca ggtgaggctt cccgcctggc gcattacaac aagcgctcga
      961 ccatcacctc cagggagatc cagacggccg tgcgcctgct gctgcctggg gagttggcca
     1021 agcacgccgt gtccgagggt actaaggcca tcaccaagta caccagcgct aaggatccag
     1081 tgagcaaggg cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc
     1141 acatggaggg ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc
     1201 cctacgaggg cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg
     1261 cctgggacat cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg
     1321 ccgacatccc cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga
     1381 tgaacttcga ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg
     1441 agttcatcta caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc
     1501 agaagaagac catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc
     1561 tgaagggcga gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg
     1621 tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca
     1681 tcaagttgga catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg
     1741 ccgagggccg ccactccacc ggcggcatgg acgagctgta ctaagctagc gcttcgaagg
     1801 cgcccgggat ccgatatcgt acgttaacat agataactga tccagtgtgc tggaattaat
     1861 tcgctgtctg cgagggccag ctgttggggt gagtactccc tctcaaaagc gggcatgact
     1921 tctgcgctaa gattgtcagt ttccaaaaac gaggaggatt tgatattcac ctggcccgcg
     1981 gtgatgcctt tgagggtggc cgcgtccatc tggtcagaaa agacaatctt tttgttgtca
     2041 agcttgaggt gtggcaggct tgagatctgg ccatacactt gagtgacaat gacatccact
     2101 ttgcctttct ctccacaggt gtccactccc aggtccaact gcaggtcgag catgcatcta
     2161 gggcggccgc actagaggaa ttccgcccct ctcccccccc cccctctccc tccccccccc
     2221 ctaacgttac tggccgaagc cgcttggaat aaggccggtg tgcgtttgtc tatatgttat
     2281 tttccaccat attgccgtct tttggcaatg tgagggcccg gaaacctggc cctgtcttct
     2341 tgacgagcat tcctaggggt ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg
     2401 tcgtgaagga agcagttcct ctggaagctt cttgaagaca aacaacgtct gtagcgaccc
     2461 tttgcaggca gcggaacccc ccacctggcg acaggtgcct ctgcggccaa aagccacgtg
     2521 tataagatac acctgcaaag gcggcacaac cccagtgcca cgttgtgagt tggatagttg
     2581 tggaaagagt caaatggctc tcctcaagcg tattcaacaa ggggctgaag gatgcccaga
     2641 aggtacccca ttgtatggga tctgatctgg ggcctcggtg cacatgcttt acatgtgttt
     2701 agtcgaggtt aaaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa
     2761 aaacacgatg ataagcttgc cacaacccgt accaaagatg gatagatccg gaaagcctga
     2821 actcaccgcg acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg tctccgacct
     2881 gatgcagctc tcggagggcg aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg
     2941 atatgtcctg cgggtaaata gctgcgccga tggtttctac aaagatcgtt atgtttatcg
     3001 gcactttgca tcggccgcgc tcccgattcc ggaagtgctt gacattgggg aattcagcga
     3061 gagcctgacc tattgcatct cccgccgtgc acagggtgtc acgttgcaag acctgcctga
     3121 aaccgaactg cccgctgttc tgcagccggt cgcggaggcc atggatgcga tcgctgcggc
     3181 cgatcttagc cagacgagcg ggttcggccc attcggaccg caaggaatcg gtcaatacac
     3241 tacatggcgt gatttcatat gcgcgattgc tgatccccat gtgtatcact ggcaaactgt
     3301 gatggacgac accgtcagtg cgtccgtcgc gcaggctctc gatgagctga tgctttgggc
     3361 cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca acaatgtcct
     3421 gacggacaat ggccgcataa cagcggtcat tgactggagc gaggcgatgt tcggggattc
     3481 ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta tggagcagca
     3541 gacgcgctac ttcgagcgga ggcatccgga gcttgcagga tcgccgcggc tccgggcgta
     3601 tatgctccgc attggtcttg accaactcta tcagagcttg gttgacggca atttcgatga
     3661 tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga tccggagccg ggactgtcgg
     3721 gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc gatggctgtg tagaagtact
     3781 cgccgatagt ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat agagtagatg
     3841 ccgaccgaac aagagctgat ttcgagaacg cctcagccag caactcgcgc gagcctagca
     3901 aggcaaatgc gagagaacgg ccttacgctt ggtggcacag ttctcgtcca cagttcgcta
     3961 agctcgctcg gctgggtcgc gggagggccg gtcgcagtga ttcaggccct tctggattgt
     4021 gttggtcccc agggcacgat tgtcatgccc acgcactcgg gtgatctgac tgatcccgca
     4081 gattggagat cgccgcccgt gcctgccgat tgggtgcaga tctctagata actgatcata
     4141 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     4201 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     4261 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4321 cattctagtt gtggtttgtc caaactcatc aatgtatctt aacgctcgag tgcattctag
     4381 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     4441 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     4501 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     4561 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     4621 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4681 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4741 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4801 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4861 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4921 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4981 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5041 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5101 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5161 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5221 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5281 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     5341 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5401 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     5461 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     5521 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     5581 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     5641 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5701 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5761 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5821 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5881 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5941 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     6001 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     6061 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     6121 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     6181 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     6241 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     6301 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     6361 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6421 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     6481 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     6541 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     6601 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc
     6661 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     6721 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     6781 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
     6841 agatatacgc gtt
//
