LOCUS       sequence-178249-        6642 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-EGFP reporter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6642)
  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 6642)
  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-GFP
FEATURES             Location/Qualifiers
     source          1..6642
                     /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             join(2824..2826,2827..2829,2830..3543)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2824..2826,2827..2829,2830..3540)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2869..2890)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3130..3149)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3477..3498
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3634..3653)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3699..3754
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3753..3772)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4145..4161)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4145..4161)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4158..4180)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4169..4185
                     /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(4193..4199,4200..4217,4218..4223))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4238..4259
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4318..4513
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4364..4499
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4519..4653
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4556..4575)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4610..4629
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4721..4738)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4892..5480)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4972..4991)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5651..6442,6443..6511))
                     /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     6274..6293
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6512..6616)
                     /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 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
     2881 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
     2941 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
     3001 accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
     3061 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     3121 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     3181 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     3241 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
     3301 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
     3361 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     3421 cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
     3481 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     3541 taagcggccg caattcactc ctcaggtgca ggctgcctat cagaaggtgg tggctggtgt
     3601 ggccaatgcc ctggctcaca aataccactg agatcttttt ccctctgcca aaaattatgg
     3661 ggacatcatg aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat
     3721 tgcaatagtg tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa
     3781 tcatttaaaa catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct
     3841 ggctgccatg aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc
     3901 tgtccattcc ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg
     3961 ttttgtgtta tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag
     4021 atttttcctc ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc
     4081 cctcgacctg caccgtcgac cagctggtcg acggtgcacc gtcgaccagc ttggcgtaat
     4141 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     4201 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     4261 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgca
     4321 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     4381 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     4441 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     4501 aggcttttgc aaaaagctaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     4561 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     4621 aaactcatca atgtatctta tcatgtctgg atccgctgca ttaatgaatc ggccaacgcg
     4681 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4741 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4801 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4861 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4921 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4981 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     5041 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta
     5101 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     5161 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     5221 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     5281 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     5341 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5401 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5461 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5521 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5581 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5641 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5701 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5761 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5821 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5881 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5941 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     6001 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6061 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6121 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6181 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6241 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6301 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6361 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6421 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6481 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6541 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6601 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tg
//
