LOCUS       sequence-471097-        6390 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Protein overexpression of cerebody.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6390)
  TITLE     A method for the detection and enrichment of endogenous cereblon 
            substrates
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6390)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pET28a-cerebody
FEATURES             Location/Qualifiers
     source          1..6390
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(778..1233)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(915..936)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1127..1146
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(1270..1317)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     1313..1331
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(1384..1401)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(2451..2474)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(2475..2501)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2502..2522)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             complement(2547..2564)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2575..2597
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    2612..2636
                     /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)."
     primer_bind     complement(2636..2655)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2637..2655)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2739..2758)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2964..3041
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3042..4124
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3061..3080)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4933..5124
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(5118..5140)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5226..5365
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5281..5300
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5380..5397)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5551..6139)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5631..5650)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(6250..6269)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             join(6261..6390,1..686)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             join(6261..6390,1..686)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(6334..6353)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg
       61 atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca
      121 tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg atccccggga
      181 aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc
      241 tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg
      301 atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga
      361 gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata
      421 aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc
      481 ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag
      541 accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac
      601 agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc
      661 atttgatgct cgatgagttt ttctaagaat taattcatga gcggatacat atttgaatgt
      721 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
      781 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
      841 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
      901 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
      961 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     1021 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     1081 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     1141 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     1201 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccaatccgg atatagttcc
     1261 tcctttcagc aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctagtta
     1321 ttgctcagcg gtggcagcag ccaactcagc ttcctttcgg gctttgttag cagccggatc
     1381 tcagtggtgg tggtggtggt gctcgagtgc ggccgcaagc ttgtcgacgg agctcgaatt
     1441 cggatccgta tccaccttta ctgttagatc gtggcaatca gcgcagagcg cgttaagccc
     1501 cagaattttt gagggctcat gtcttttttg gtggccgtaa acttccatcc aatatggctc
     1561 gcacagatct tgcactgggc aaccgtccag gcataccctg gaaaccagct gtgttcggta
     1621 gacgggcggc ctatcaggtt caggttgcac gccttataca cagtcagaat ctcatgcaca
     1681 tacccatgcg gattcacata cgctgccatc ggaccctcgc gggacagcga gaaaatttcg
     1741 tttttggttg tgatttcggt ttcttgacat tgtttgcagc acagggaagt acatttattc
     1801 ataatgtcta attcacagcg aagtcgctgg atcgccgaat caattttcag gagctgaata
     1861 cgcaagctat catcaatagg aaggttcgcc gctacccagt agctaaaatc aattggattt
     1921 gaaggcagcg aatcatcttt gagattctca tcccattcac ggagctgttt cttgatacgg
     1981 tccattaagg tctccgcatc gtcatagcgt tccggaagaa tctgcacttt agcttgctgg
     2041 ataccatctg actgggtgcg tagctcaagg actttgaaac gttgacggcc aattgcttta
     2101 actttcacga tctcaatacc gaaatcctgt tcttcacgat aggcgtaaat ctcggctgta
     2161 gtgccaaatt ctgcttccct ttcctgtaca ttgctgtatg ctaaaacagc aaaggtgcga
     2221 tcgttctgaa tcagattccg gaccatactg acttcctgcg ggtgaaacag ctgcagcggt
     2281 aaggtctgtc cgggaaccag gatcatcatc acttgcggca gaaccggaat cacctgaatg
     2341 ctgtcgtcat cgtgcaacgt gcggccatga aattcttcca tatcggcacc caggtaggtg
     2401 tgtgatgtcg gcagactggt gtcgaagttg atgatgtttg gctttttggc cttgtcatcg
     2461 tcatctttat aatcagcgta atctggaaca tcgtatgggt aggattggaa gtacaggttc
     2521 tcggtaccca gatctacacc agaagaatga tgatgatgat ggtgcatatg tatatctcct
     2581 tcttaaagtt aaacaaaatt atttctagag gggaattgtt atccgctcac aattccccta
     2641 tagtgagtcg tattaatttc gcgggatcga gatctcgatc ctctacgccg gacgcatcgt
     2701 ggccggcatc accggcgcca caggtgcggt tgctggcgcc tatatcgccg acatcaccga
     2761 tggggaagat cgggctcgcc acttcgggct catgagcgct tgtttcggcg tgggtatggt
     2821 ggcaggcccc gtggccgggg gactgttggg cgccatctcc ttgcatgcac cattccttgc
     2881 ggcggcggtg ctcaacggcc tcaacctact actgggctgc ttcctaatgc aggagtcgca
     2941 taagggagag cgtcgagatc ccggacacca tcgaatggcg caaaaccttt cgcggtatgg
     3001 catgatagcg cccggaagag agtcaattca gggtggtgaa tgtgaaacca gtaacgttat
     3061 acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg
     3121 ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt
     3181 acattcccaa ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg attggcgttg
     3241 ccacctccag tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg
     3301 ccgatcaact gggtgccagc gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct
     3361 gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc attaactatc
     3421 cgctggatga ccaggatgcc attgctgtgg aagctgcctg cactaatgtt ccggcgttat
     3481 ttcttgatgt ctctgaccag acacccatca acagtattat tttctcccat gaagacggta
     3541 cgcgactggg cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg
     3601 gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa tatctcactc
     3661 gcaatcaaat tcagccgata gcggaacggg aaggcgactg gagtgccatg tccggttttc
     3721 aacaaaccat gcaaatgctg aatgagggca tcgttcccac tgcgatgctg gttgccaacg
     3781 atcagatggc gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg
     3841 atatctcggt agtgggatac gacgataccg aagacagctc atgttatatc ccgccgttaa
     3901 ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac
     3961 tctctcaggg ccaggcggtg aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa
     4021 aaaccaccct ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     4081 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     4141 gtaagttagc tcactcatta ggcaccggga tctcgaccga tgcccttgag agccttcaac
     4201 ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact tatgactgtc
     4261 ttctttatca tgcaactcgt aggacaggtg ccggcagcgc tctgggtcat tttcggcgag
     4321 gaccgctttc gctggagcgc gacgatgatc ggcctgtcgc ttgcggtatt cggaatcttg
     4381 cacgccctcg ctcaagcctt cgtcactggt cccgccacca aacgtttcgg cgagaagcag
     4441 gccattatcg ccggcatggc ggccccacgg gtgcgcatga tcgtgctcct gtcgttgagg
     4501 acccggctag gctggcgggg ttgccttact ggttagcaga atgaatcacc gatacgcgag
     4561 cgaacgtgaa gcgactgctg ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc
     4621 ttcggtttcc gtgtttcgta aagtctggaa acgcggaagt cagcgccctg caccattatg
     4681 ttccggatct gcatcgcagg atgctgctgg ctaccctgtg gaacacctac atctgtatta
     4741 acgaagcgct ggcattgacc ctgagtgatt tttctctggt cccgccgcat ccataccgcc
     4801 agttgtttac cctcacaacg ttccagtaac cgggcatgtt catcatcagt aacccgtatc
     4861 gtgagcatcc tctctcgttt catcggtatc attaccccca tgaacagaaa tcccccttac
     4921 acggaggcat cagtgaccaa acaggaaaaa accgccctta acatggcccg ctttatcaga
     4981 agccagacat taacgcttct ggagaaactc aacgagctgg acgcggatga acaggcagac
     5041 atctgtgaat cgcttcacga ccacgctgat gagctttacc gcagctgcct cgcgcgtttc
     5101 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     5161 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     5221 cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtatactgg cttaactatg
     5281 cggcatcaga gcagattgta ctgagagtgc accattgcgg tgtgaaatac cgcacagatg
     5341 cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5401 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5461 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5521 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5581 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5641 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5701 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5761 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5821 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5881 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5941 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt
     6001 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6061 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6121 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6181 gaacgaaaac tcacgttaag ggattttggt catgaacaat aaaactgtct gcttacataa
     6241 acagtaatac aaggggtgtt atgagccata ttcaacggga aacgtcttgc tctaggccgc
     6301 gattaaattc caacatggat gctgatttat atgggtataa atgggctcgc gataatgtcg
     6361 ggcaatcagg tgcgacaatc tatcgattgt
//
