LOCUS       sequence-83952-e        5260 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Contains human Histone H2B fused to EGFP in the pCS2+ expression 
            vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5260)
  AUTHORS   Megason SG
  TITLE     In toto imaging of embryogenesis with confocal time-lapse 
            microscopy.
  JOURNAL   Methods Mol Biol. 2009;546:317-32. doi: 
            10.1007/978-1-60327-977-2_19.
  PUBMED    19378112
REFERENCE   2  (bases 1 to 5260)
  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: "Methods Mol
            Biol."; date: "2009"; volume: "546:317-32. doi"; pages: " 
            10.1007/978-1-60327-977-2_19"
COMMENT     Sequence Label: pCS-H2B-EGFP
FEATURES             Location/Qualifiers
     source          1..5260
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(45..65)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(45..63)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(84..100)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(84..100)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(97..119)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    108..124
                     /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(132..138,139..156,157..162))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    177..198
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(315..332)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(486..1074)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(566..585)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1245..2036,2037..2105))
                     /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     1868..1887
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2106..2210)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2236..2691)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2373..2394)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2585..2604
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2765..3749
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        3784..3802
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3784..3801
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             3907..4284
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     regulatory      4297..4306
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4303..4305,4306..4308,4309..5022)
                     /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(4303..4305,4306..4308,4309..5019)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4348..4369)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4609..4628)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4956..4977
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5058..5192
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5082..5101)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5136..5155
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
ORIGIN
        1 ggccgcggcg ccaatgcatt gggcccggta cccagctttt gttcccttta gtgagggtta
       61 attgcgcgct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
      121 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
      181 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
      241 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
      301 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
      361 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
      421 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
      481 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
      541 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
      601 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
      661 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
      721 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
      781 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
      841 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
      901 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
      961 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     1021 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     1081 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     1141 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     1201 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     1261 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     1321 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     1381 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     1441 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     1501 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     1561 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     1621 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     1681 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     1741 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     1801 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     1861 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     1921 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     1981 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     2041 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     2101 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     2161 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     2221 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     2281 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     2341 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     2401 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     2461 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     2521 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     2581 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     2641 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc
     2701 cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc
     2761 agtcgaccat agccaattca atatggcgta tatggactca tgccaattca atatggtgga
     2821 tctggacctg tgccaattca atatggcgta tatggactcg tgccaattca atatggtgga
     2881 tctggacccc agccaattca atatggcgga cttggcacca tgccaattca atatggcgga
     2941 cttggcactg tgccaactgg ggaggggtct acttggcacg gtgccaagtt tgaggagggg
     3001 tcttggccct gtgccaagtc cgccatattg aattggcatg gtgccaataa tggcggccat
     3061 attggctata tgccaggatc aatatatagg caatatccaa tatggcccta tgccaatatg
     3121 gctattggcc aggttcaata ctatgtattg gccctatgcc atatagtatt ccatatatgg
     3181 gttttcctat tgacgtagat agcccctccc aatgggcggt cccatatacc atatatgggg
     3241 cttcctaata ccgcccatag ccactccccc attgacgtca atggtctcta tatatggtct
     3301 ttcctattga cgtcatatgg gcggtcctat tgacgtatat ggcgcctccc ccattgacgt
     3361 caattacggt aaatggcccg cctggctcaa tgcccattga cgtcaatagg accacccacc
     3421 attgacgtca atgggatggc tcattgccca ttcatatccg ttctcacgcc ccctattgac
     3481 gtcaatgacg gtaaatggcc cacttggcag tacatcaata tctattaata gtaacttggc
     3541 aagtacatta ctattggaag gacgccaggg tacattggca gtactcccat tgacgtcaat
     3601 ggcggtaaat ggcccgcgat ggctgccaag tacatcccca ttgacgtcaa tggggagggg
     3661 caatgacgca aatgggcgtt ccattgacgt aaatgggcgg taggcgtgcc taatgggagg
     3721 tctatataag caatgctcgt ttagggaacc gccattctgc ctggggacgt cggagcaagc
     3781 ttgatttagg tgacactata gaatacaagc tacttgttct ttttgcagga tcccatcgat
     3841 tcgaattcaa ggcctctcga gcctctagaa ctatagtgag tcgtattact cgacggtacc
     3901 gccaccatgc cagagccagc gaagtctgct cccgccccga aaaagggctc caagaaggcg
     3961 gtgactaagg cgcagaagaa aggcggcaag aagcgcaagc gcagccgcaa ggagagctat
     4021 tccatctatg tgtacaaggt tctgaagcag gtccaccctg acaccggcat ttcgtccaag
     4081 gccatgggca tcatgaattc gtttgtgaac gacattttcg agcgcatcgc aggtgaggct
     4141 tcccgcctgg cgcattacaa caagcgctcg accatcacct ccagggagat ccagacggcc
     4201 gtgcgcctgc tgctgcctgg ggagttggcc aagcacgccg tgtccgaggg tactaaggcc
     4261 atcaccaagt acaccagcgc taaggatcca ccggtcgcca ccatggtgag caagggcgag
     4321 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     4381 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     4441 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
     4501 tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag
     4561 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     4621 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     4681 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     4741 aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc
     4801 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
     4861 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     4921 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     4981 gccgccggga tcactctcgg catggacgag ctgtacaagt aaagcggccc tagaactata
     5041 gtgagtcgta ttacgtagat ccagacatga taagatacat tgatgagttt ggacaaacca
     5101 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
     5161 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt
     5221 ttcaggttca gggggaggtg tgggaggttt tttaattcgc
//
