LOCUS       sequence-474676-        5266 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express LgBiT-CALM1 Fusion Protein in Mammalian Cells under a CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5266)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5266)
  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: 
            "Unpublished"
COMMENT     Sequence Label: LgBiT-CALM1 BiBiT-Ready Vector
FEATURES             Location/Qualifiers
     source          1..5266
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     573..592
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(861..1449)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(941..960)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    1575..1623
                     /note="synthetic polyadenylation signal"
     CDS             complement(1647..2684)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1660..1668
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(2715..3072)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2729..2864
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        2881..3072
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     3029..3049
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    3144..3225
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     3169..3188
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3428..3877)
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="MADQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPT
                     EAELQDMINEVDADGNGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYISAA
                     ELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     promoter        complement(4447..4465)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4448..4465)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        4585..4888
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5043..5063
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        5096..5143
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             join(5175..5266,1..703)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5229..5248)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
       61 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
      121 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
      181 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
      241 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
      301 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
      361 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
      421 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
      481 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
      541 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
      601 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
      661 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgattcgaaa tgaccgacca
      721 agcgacgccc aaccggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
      781 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
      841 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
      901 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
      961 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     1021 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     1081 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     1141 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     1201 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     1261 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     1321 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     1381 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     1441 ggatttcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     1501 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     1561 tagtccggaa tcgtcacaca aaaaaccaac acacagatgt aatgaaaata aagatatttt
     1621 attgcggcca tcgtgatggc tagagtctaa actcgaccta cctccttagc gcgcggtcgt
     1681 gtactgggcc ggcggttgcc gctgtcggcc agcacctcga cgcagccgtc ggtccatacg
     1741 gctgcgctcc ggcgagcgat ttgtgtgcga ccgacggtgc cggccccgct gcggacgatg
     1801 gcatcgcagc ggccttgagc ccaggcagca tcgtcgaagt tgccgtccac gaggctctgg
     1861 tagagctgat ccaggccgat gcgcagcatg taggctcgca gacgagggct gccggccagc
     1921 tcgggatgcc ggcgctcgaa gtagcgagtc tgctgctcca tgcaagccag ccagggccgc
     1981 cagaagaaga tgttggccac ctcgtactga ctgtccccga acatagcttc ggaccagtcg
     2041 attacggcgg tgatgcggcc gttgtcggtc aggacgttgt tgctgccgaa gtcggcatgg
     2101 acgaggtggc gcacctcggg acagtcttcg gcccacagca tcagttcgtc cagggcttga
     2161 gctacgctgg cggacacggt gtcgtccatc acggtctgcc agtggtagac atggggatca
     2221 gcaatggcgc aaatgaaatc ccgccaggtt gtgtactggc cgatgccttg gggcccgaat
     2281 gggccgaagc cgctggtttg actgaggtcg gcggcggcaa tagcatccat agcttcggcg
     2341 acaggctgta acacagcagg cagctctgtt tcggggaggt cttggagagt aacgccttgg
     2401 gcgcgtctac tgatgcagta tgtcaggctc tcgctgaact cgccgatgtc caacacttcg
     2461 gggatgggta gtgcagcgct ggcgaagtgg cggtacacgt agcggtcttt gtagaagcca
     2521 tcagcgcagc tattcacccg cagtacatag ccgcgtccgc cgacatcgaa gctgaaggct
     2581 cggctctctt cgccctccga caactgcatc aggtcgctca cactgtcgaa cttctcgatg
     2641 agaaattttt caacgctggt agcggtgagt tcgggcttct tcatggtggc gctagtgtca
     2701 gaagaatcga gctttttgca aaagcctagg cctccaaaaa agcctcctca ctacttctgg
     2761 aatagctcag aggccgaggc ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca
     2821 tggggcggag aatgggcgga actgggcgga gttaggggcg ggatgggcgg agttaggggc
     2881 gggactatgg ttgctgacta attgagatgc atgctttgca tacttctgcc tgctggggag
     2941 cctggggact ttccacacct ggttgctgac taattgagat gcatgctttg catacttctg
     3001 cctgctgggg agcctgggga ctttccacac cctaactgac acacattcca cagctggttc
     3061 tttccgcctc agaaggtacc taaccaagtt cctctttcag aggttatttc aggccatggt
     3121 gctgcgcaag acgcgtgacg tcggcagtga aaaaaatgct ttatttgtga aatttgtgat
     3181 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     3241 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
     3301 ctctacaaat gtggtatggc tgattatgat ccgcggccgc aatacgtcga cgttatcagc
     3361 tgacttcgta cgagagccta ggtgtttcta gaggatcccc gggtaccgag cccgaattcg
     3421 tttaaactca ttttgcagtc atcatctgta cgaattcttc atagttgact tgtccgtctc
     3481 catcaatatc tgcttctctg atcatttcat ctacttcttc atctgttagt ttttctccta
     3541 agtttgtcat gacgtgacgt agttctgctg cactgatata accattgcca tccttgtcaa
     3601 agactcggaa tgcctcacgg atttcttctt cactatctgt atctttcatt tttctagcca
     3661 tcatagtcaa aaattcgggg aagtcaatgg tgccattacc atcagcatcc acttcattga
     3721 tcatatcctg caattcagct tctgttgggt tctgacccag tgacctcatg acagttccaa
     3781 gttcctttgt tgtgatggtg ccatcgccat ctttatcaaa tagggagaag gcttccttga
     3841 attcagcaat ctgttcttcg gtcagctgat cagccatggc gatcgcaccg ctcgagcctc
     3901 cacctccgct cccgccacca ccggagctcc cactgttgat ggttactcgg aacagcatgg
     3961 agccgtcggg ggtgatcagg cgctcgtcga taattttgtt gccgttccac agggtccctg
     4021 ttacagtgat ctttttgccg tcgaacacgg cgatgccttc atacggccgt ccgaaatagt
     4081 tcagcatgtt cggcgtaacc ccgtcgatta ccagtgtgcc atagggcagg atcaccttaa
     4141 agtgatgatc atccacaggg tacaccacct taaacacctc ttcgatctgg gccatttggt
     4201 cggcgctcag accttcatac gggatgatga catggatgtc gatcttcagg gcattttcac
     4261 cgctccggac aatcctttgg atcggagtta cggacacggc gagattctgc agcaaactgg
     4321 acacacctcc ctgttcaagg acttggtcca ggttgtaggc ggctgtctgt tcccagtccc
     4381 caacgaaatc ttcgagtgtg aagaccatgg tgagcgctag ctgcaaaaag aacaagtagc
     4441 ttgtattcta tagtgtcacc taaatgggcg atctgacggt tcactaaacc agctctgctt
     4501 atataggcct ccctggcgcg ccactagtta ttaatagtaa tcaattacgg ggtcattagt
     4561 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     4621 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     4681 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     4741 agtacatcaa gtgtatcata tgccaagtcc gccccctatt gacgtcaatg acggtaaatg
     4801 gcccgcctgg cattatgccc agtacatgac cttacgggac tttcctactt ggcagtacat
     4861 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca ccaatgggcg
     4921 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     4981 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaataacc ccgccccgtt
     5041 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtcaaa tcgaattcta atactgttga
     5101 caattaatca tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac
     5161 ccaggaggca gatcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     5221 tggagaggct attcggctat gactgggcac aacagacaat cggctg
//
