LOCUS       sequence-178236-        6600 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ubiquitous expression plasmid, contains CAG promoter (CMV immediate 
            early enhancer and chicken beta actin promoter), MCS and IRES 
            controlled Histone2B-mRFP1 reporter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6600)
  AUTHORS   Williams RM, Senanayake U, Artibani M, Taylor G, Wells D, Ahmed AA, 
            Sauka-Spengler T
  TITLE     Genome and epigenome engineering CRISPR toolkit for in vivo 
            modulation of cis-regulatory interactions and gene expression in the
            chicken embryo.
  JOURNAL   Development. 2018 Feb 23;145(4). pii: dev.160333. doi: 
            10.1242/dev.160333.
  PUBMED    29386245
REFERENCE   2  (bases 1 to 6600)
  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: 
            "Development."; date: "2018-02-23"; volume: "145(4). pii: 
            dev.160333. doi"; pages: " 10.1242/dev.160333"
COMMENT     Sequence Label: pCI H2B-RFP
FEATURES             Location/Qualifiers
     source          1..6600
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1674
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1598..1620
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1682..1701
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     misc_feature    join(1835..2409,2410..2412,2413..2421)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2002..2019)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2229..2248
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2428..2805
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     regulatory      2818..2827
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2824..3501
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             2824..3501
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(2954..2972)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3222..3241
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3416..3439
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(3592..3611)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3657..3712
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3711..3730)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4103..4119)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4103..4119)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4116..4138)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4127..4143
                     /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(4151..4157,4158..4175,4176..4181))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4196..4217
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4276..4471
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4322..4457
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4477..4611
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4514..4533)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4568..4587
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4679..4696)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4850..5438)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4930..4949)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5609..6400,6401..6469))
                     /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     6232..6251
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6470..6574)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtac ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gcccccggag cgccggcggc
     1321 tgtcgaggcg cggcgagccg cagccattgc cttttatggt aatcgtgcga gagggcgcag
     1381 ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc
     1441 tctagcgggc gcggggcgaa gcggtgcggc gccggcagga aggaaatggg cggggagggc
     1501 cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1681 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattgct cgagagttcg
     1741 aagccttaag tacgcgtagg cgcgccttaa ttaacgttta aacgctagct catcgataag
     1801 cttgatatcg aattcctgca gcccggggga tccgcccctc tccctccccc ccccctaacg
     1861 ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca
     1921 ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc ttcttgacga
     1981 gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg aatgtcgtga
     2041 aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg accctttgca
     2101 ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag
     2161 atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata gttgtggaaa
     2221 gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc cagaaggtac
     2281 cccattgtat gggatctgat ctggggcctc ggtacacatg ctttacatgt gtttagtcga
     2341 ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt tgaaaaacac
     2401 gatgataata tggccacaac catggttatg ccagagccag cgaagtctgc tcccgccccg
     2461 aaaaagggct ccaagaaggc ggtgactaag gcgcagaaga aaggcggcaa gaagcgcaag
     2521 cgcagccgca aggagagcta ttccatctat gtgtacaagg ttctgaagca ggtccaccct
     2581 gacaccggca tttcgtccaa ggccatgggc atcatgaatt cgtttgtgaa cgacattttc
     2641 gagcgcatcg caggtgaggc ttcccgcctg gcgcattaca acaagcgctc gaccatcacc
     2701 tccagggaga tccagacggc cgtgcgcctg ctgctgcctg gggagttggc caagcacgcc
     2761 gtgtccgagg gtactaaggc catcaccaag tacaccagcg ctaaggatcc accggtcgcc
     2821 accatggcct cctccgagga cgtcatcaag gagttcatgc gcttcaaggt gcgcatggag
     2881 ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag
     2941 ggcacccaga ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac
     3001 atcctgtccc ctcagttcca gtacggctcc aaggcctacg tgaagcaccc cgccgacatc
     3061 cccgactact tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc
     3121 gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc
     3181 tacaaggtga agctgcgcgg caccaacttc ccctccgacg gccccgtaat gcagaagaag
     3241 accatgggct gggaggcctc caccgagcgg atgtaccccg aggacggcgc cctgaagggc
     3301 gagatcaaga tgaggctgaa gctgaaggac ggcggccact acgacgccga ggtcaagacc
     3361 acctacatgg ccaagaagcc cgtgcagctg cccggcgcct acaagaccga catcaagctg
     3421 gacatcacct cccacaacga ggactacacc atcgtggaac agtacgagcg cgccgagggc
     3481 cgccactcca ccggcgccta agcggccgca attcactcct caggtgcagg ctgcctatca
     3541 gaaggtggtg gctggtgtgg ccaatgccct ggctcacaaa taccactgag atctttttcc
     3601 ctctgccaaa aattatgggg acatcatgaa gccccttgag catctgactt ctggctaata
     3661 aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct cactcggaag
     3721 gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt agagtttggc
     3781 aacatatgcc catatgctgg ctgccatgaa caaaggttgg ctataaagag gtcatcagta
     3841 tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga cttgaggtta
     3901 gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa aattttcctt
     3961 acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca tagctgtccc
     4021 tcttctctta tggagatccc tcgacctgca ccgtcgacca gctggtcgac ggtgcaccgt
     4081 cgaccagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4141 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4201 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4261 cgtgccagcg gatccgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     4321 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     4381 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     4441 aggctttttt ggaggcctag gcttttgcaa aaagctaact tgtttattgc agcttataat
     4501 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     4561 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggat ccgctgcatt
     4621 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     4681 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4741 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4801 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4861 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4921 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4981 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5041 ctcaatgctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5101 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5161 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5221 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5281 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5341 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5401 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5461 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5521 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     5581 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5641 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5701 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     5761 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5821 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5881 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     5941 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6001 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6061 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6121 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6181 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6241 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6301 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6361 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6421 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6481 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6541 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
//
