LOCUS       sequence-353789-        6783 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for the purpose of protein purification..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6783)
  AUTHORS   Modzelewski AJ, Shao W, Chen J, Lee A, Qi X, Noon M, Tjokro K, Sales
            G, Biton A, Anand A, Speed TP, Xuan Z, Wang T, Risso D, He L
  TITLE     A mouse-specific retrotransposon drives a conserved Cdk2ap1 isoform 
            essential for development.
  JOURNAL   Cell. 2021 Oct 7. pii: S0092-8674(21)01104-1. doi: 
            10.1016/j.cell.2021.09.021.
  PUBMED    34644528
REFERENCE   2  (bases 1 to 6783)
  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: "Cell. 2021 
            Oct 7. pii: S0092-8674(21)01104-1. doi: 10.1016/j.cell.2021.09.021."
COMMENT     Sequence Label: pET28a Cdk2ap1-Delta-N (MT2B2)
FEATURES             Location/Qualifiers
     source          1..6783
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(50..865)
                     /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             complement(50..865)
                     /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     773..792
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     857..876
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      987..1575
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1476..1495
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1729..1746
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1761..1900
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1826..1845)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1986..2008
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2002..2193)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3002..4084)
                     /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     4046..4065
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4085..4162)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     4368..4387
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4471..4490
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4471..4489
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4490..4514
                     /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)."
     RBS             4529..4551
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4570..4587
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4597..4614
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             4618..5718
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      5575..5584
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     5692..5715
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             5734..5754
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             5806..5826
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             6118..6135
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6188..6206)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6202..6249
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      6286..6741
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6373..6392)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6583..6604
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 tttatttttc taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact
       61 catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt
      121 gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa
      181 gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc
      241 cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg
      301 agaatggcaa aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct
      361 cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga
      421 gacgaaatac gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc
      481 gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata
      541 cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac
      601 ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca
      661 tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg
      721 catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag
      781 cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag
      841 acgtttcccg ttgaatatgg ctcataacac cccttgtatt actgtttatg taagcagaca
      901 gttttattgt tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
      961 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     1021 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     1081 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     1141 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     1201 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     1261 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     1321 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     1381 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     1441 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     1501 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     1561 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     1621 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     1681 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     1741 agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     1801 tcacaccgca atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     1861 atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc
     1921 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     1981 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca
     2041 gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc
     2101 cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc
     2161 catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct
     2221 gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga
     2281 tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg
     2341 gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg
     2401 tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg
     2461 cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca ttcatgttgt
     2521 tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga
     2581 ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag
     2641 cacgatcatg cgcacccgtg gggccgccat gccggcgata atggcctgct tctcgccgaa
     2701 acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac
     2761 cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac
     2821 ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag tcataagtgc
     2881 ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa
     2941 gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg
     3001 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     3061 acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg
     3121 agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt
     3181 ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat
     3241 aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca ccaacgcgca
     3301 gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca
     3361 tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg
     3421 cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat
     3481 gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga
     3541 tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg
     3601 agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat
     3661 tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca
     3721 gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc
     3781 ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg
     3841 ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca
     3901 acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca
     3961 tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc
     4021 gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt
     4081 tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg
     4141 ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt
     4201 aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca
     4261 tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg
     4321 aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
     4381 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
     4441 tagaggatcg agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag
     4501 cggataacaa ttcccctcta gaaataattt tgtttaactt taagaaggag atataccatg
     4561 ggcagcagcc atcatcatca tcatcacagc agcggcctgg tgccgcgcgg cagccatatg
     4621 aaaatcgaag aaggtaaact ggtaatctgg attaacggcg ataaaggcta taacggtctc
     4681 gctgaagtcg gtaagaaatt cgagaaagat accggaatta aagtcaccgt tgagcatccg
     4741 gataaactgg aagagaaatt cccacaggtt gcggcaactg gcgatggccc tgacattatc
     4801 ttctgggcac acgaccgctt tggtggctac gctcaatctg gcctgttggc tgaaatcacc
     4861 ccggacaaag cgttccagga caagctgtat ccgtttacct gggatgccgt acgttacaac
     4921 ggcaagctga ttgcttaccc gatcgctgtt gaagcgttat cgctgattta taacaaagat
     4981 ctgctgccga acccgccaaa aacctgggaa gagatcccgg cgctggataa agaactgaaa
     5041 gcgaaaggta agagcgcgct gatgttcaac ctgcaagaac cgtacttcac ctggccgctg
     5101 attgctgctg acgggggtta tgcgttcaag tatgaaaacg gcaagtacga cattaaagac
     5161 gtgggcgtgg ataacgctgg cgcgaaagcg ggtctgacct tcctggttga cctgattaaa
     5221 aacaaacaca tgaatgcaga caccgattac tccatcgcag aagctgcctt taataaaggc
     5281 gaaacagcga tgaccatcaa cggcccgtgg gcatggtcca acatcgacac cagcaaagtg
     5341 aattatggtg taacggtact gccgaccttc aagggtcaac catccaaacc gttcgttggc
     5401 gtgctgagcg caggtattaa cgccgccagt ccgaacaaag agctggcaaa agagttcctc
     5461 gaaaactatc tgctgactga tgaaggtctg gaagcggtta ataaagacaa accgctgggt
     5521 gccgtagcgc tgaagtctta cgaggaagag ttggcgaaag atccacgtat tgccgccacc
     5581 atggaaaacg cccagaaagg tgaaatcatg ccgaacatcc cgcagatgtc cgctttctgg
     5641 tatgccgtgc gtactgcggt gatcaacgcc gccagcggtc gtcagactgt cgatgaagcc
     5701 ctgaaagacg cgcagactaa ttcgaatttc gaggaaaatt tatattttca aagcggcacc
     5761 gtggatgccg gcgccgatgc cggcaagccc cggcccggca gcagcgagaa cctgtacttc
     5821 cagggcggcc ggccgatggc gacctcctcc caatatcgcc agctgctgag tgactatggg
     5881 cctccatcgc taggctacac ccagggaact ggaaatagcc aggtgcctca gagtaaatat
     5941 gcagagctgc tggccatcat tgaagagttg gggaaagaga tcagaccgac gtatgcaggg
     6001 agcaagagcg ccatggagag gctaaaacga ggcatcattc atgcccggag cctggttcgg
     6061 gagtgcttgg ctgaaacgga acggaatgcc aggtcctgat gagcggccgc actcgagcac
     6121 caccaccacc accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct
     6181 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     6241 ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg ccctgtagcg
     6301 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     6361 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     6421 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     6481 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     6541 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     6601 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     6661 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     6721 aaatattaac gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat
     6781 ttg
//
