LOCUS       sequence-345741-        7671 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  cDNA of CER11/CPL2 cloned into pET28a for protein expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7671)
  AUTHORS   Shi L, Dean GH, Zheng H, Meents MJ, Haslam TM, Haughn GW, Kunst L
  TITLE     ECERIFERUM11/C-TERMINAL DOMAIN PHOSPHATASE-LIKE2 Affects Secretory 
            Trafficking.
  JOURNAL   Plant Physiol. 2019 Nov;181(3):901-915. doi: 10.1104/pp.19.00722. 
            Epub 2019 Sep 4.
  PUBMED    31484679
REFERENCE   2  (bases 1 to 7671)
  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"; doi: 
            "10.1104/pp.19.00722"; journalName: "Plant Physiol"; date: 
            "2019-11"; volume: "181"; issue: "3"; pages: "901-915"
COMMENT     Sequence Label: 6xHis-CPL2/pET28a
FEATURES             Location/Qualifiers
     source          1..7671
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(2513..2545)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             complement(2549..2566)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(2576..2593)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2612..2634
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    2649..2673
                     /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(2673..2692)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2674..2692)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2776..2795)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        3001..3078
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3079..4161
                     /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(3098..3117)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4970..5161
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(5155..5177)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5263..5402
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5318..5337
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5417..5434)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5588..6176)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5668..5687)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(6287..6306)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             6298..7113
                     /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             6298..7113
                     /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(6371..6390)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(7205..7660)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7342..7363)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7554..7573
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgactcacaa ctctgaaatg ttttctttag gcgcttcctc ttcaagtcct ttgtttgata
      241 catcttcact gctttcccac aaaaatccat tatcattttc cggacctttc tctgtttctc
      301 tagctagagg tgccacatgt gcaacatact tttcagccaa acttcgaaga gcattctcag
      361 ccgcttgttg atgtgcatct ttcttagttt ttgccatccc aatgcctatc ttctcacccg
      421 tgaacaacac ctcaactgaa aactgtaact ctttgttggt acttatcaca gtcctaaact
      481 ccacctttga accacatctt cttccaattt cttgcagtgc actcacaaat aaatgagatt
      541 gcccgccatt actctttcca ccatctgtat gatgttctct accatttgac accaaatgat
      601 tctgggatat tccaccttct gttgtttgac gtgaaggatt ttgccttttc aggtcatcat
      661 ccattgctac ctcttcactc ctcaagtggg atggatgtgc aaatggacca acaggagcag
      721 aacctggtgt tccatgagga aattgacttg gataaagtcc tgaaggccga ccaggaaaag
      781 aaggtctatt ctcatcatct accaaccaac cacccggaga atgcatagac gaggaaggtg
      841 gctgcatagg aattttagcc agaatggggg gttgattaaa gttttggttc cttatatcta
      901 caccaggcct catcataagg ggtcttccac catcagaaaa agtgaagcca tcccgcctag
      961 gagctccaag caaactaggt ttatgagaag gtaacagtgc tgctgcagtt gccgtgcttg
     1021 cattatttgg tataactgct atctgcggct taggggtttc aggtttctgc tcagcgttac
     1081 ttgttgcagg aagagtagaa tgatcagctg cagccgcttg gtttagtctt cgttccactt
     1141 cacccccaca cattccttcg ttaattggag gagcatttat gtttccattt gaagcaaagc
     1201 caggatcctc tataacgacg tagttgctca catccggtga tggaggtaaa ttttccacat
     1261 catcttcata gtaaaccaaa gatatactac tcattaaact ctcatcaaat tccttgaaaa
     1321 agtaacctct gacattgcat gcaacatttc ttgctacgca gagatgtgga acgacaaggg
     1381 ctgtctctgc ttgtggagca taataaggaa gatatgcaga aacaacatga acccgtggtt
     1441 gatccttgtc ctcccaaacc ttcatacggt catcaatgac cattgccatt ttcgggtggc
     1501 aaatgccacc attgaagaca cttaacaacg atttcttggc gtctggtttt acgcagacga
     1561 tacgatctcg aagctccttg agacttatca agtgtgcttc cggatcaagc agtctccaca
     1621 tttccagagc ataatccctt tcagccatgg tacaaacgta aacttcaaaa cgtttcctag
     1681 ttttggctgt caaataactt cttaactcct cccatgctgg tctcagtttt acaagcacac
     1741 tagtgtcacg gatctctggc ttaatccgag taagcacagt attcttctcc ggcaatctaa
     1801 tcacgggtct acaaactttc tcttgtccat cagaagttgg gcggacttcc tcaggttgag
     1861 cctttaacaa aactccgtta tcaaaagcat agtcattatc aatatactgc ttcagcagca
     1921 tcctatcatc catatacctt ttaagttctg ctgacatccc attaatcctc acaggatcca
     1981 tctcacgtga aatccaagat ttaagagcct cgattctatc ctcaaatgac ttcatagtat
     2041 ttgcaacaat caaagtctca tccaaatcaa acacaataga caaacaccga gtattaagca
     2101 tccttaagca cgaatcataa agacctgaag gaaccgaaaa acaccaaaaa cacgggaact
     2161 tcttctcctt actcggcatc gccactaaat gaatctcctc atctccgagc ataaccactg
     2221 cagtcttcaa ttcatggaaa cagactgcgt gtagatgcat taactcttga gatttcaccg
     2281 gagccgatga ctcaagcttg catcgaacgg caaaagaagc aatcgtctga agaatcgcga
     2341 gcggaggaca tctctcacca gccggagata gatggtggat tcgaatctca tcgttcggga
     2401 atcgaaattc atgacttgaa gatacatggt tcacatcaag ttcccccaaa cgtaaatctc
     2461 cgtgataaac cacggattta tgacccaaac gattcatgaa ttcggatccg cgacccattt
     2521 gctgtccacc agtcatgcta gccatatggc tgccgcgcgg caccaggccg ctgctgtgat
     2581 gatgatgatg atggctgctg cccatggtat atctccttct taaagttaaa caaaattatt
     2641 tctagagggg aattgttatc cgctcacaat tcccctatag tgagtcgtat taatttcgcg
     2701 ggatcgagat ctcgatcctc tacgccggac gcatcgtggc cggcatcacc ggcgccacag
     2761 gtgcggttgc tggcgcctat atcgccgaca tcaccgatgg ggaagatcgg gctcgccact
     2821 tcgggctcat gagcgcttgt ttcggcgtgg gtatggtggc aggccccgtg gccgggggac
     2881 tgttgggcgc catctccttg catgcaccat tccttgcggc ggcggtgctc aacggcctca
     2941 acctactact gggctgcttc ctaatgcagg agtcgcataa gggagagcgt cgagatcccg
     3001 gacaccatcg aatggcgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt
     3061 caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt
     3121 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
     3181 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
     3241 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
     3301 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
     3361 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
     3421 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
     3481 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca
     3541 cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg
     3601 gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg
     3661 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
     3721 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
     3781 gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
     3841 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
     3901 gataccgaag acagctcatg ttatatcccg ccgttaacca ccatcaaaca ggattttcgc
     3961 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
     4021 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
     4081 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     4141 cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagctca ctcattaggc
     4201 accgggatct cgaccgatgc ccttgagagc cttcaaccca gtcagctcct tccggtgggc
     4261 gcggggcatg actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg
     4321 acaggtgccg gcagcgctct gggtcatttt cggcgaggac cgctttcgct ggagcgcgac
     4381 gatgatcggc ctgtcgcttg cggtattcgg aatcttgcac gccctcgctc aagccttcgt
     4441 cactggtccc gccaccaaac gtttcggcga gaagcaggcc attatcgccg gcatggcggc
     4501 cccacgggtg cgcatgatcg tgctcctgtc gttgaggacc cggctaggct ggcggggttg
     4561 ccttactggt tagcagaatg aatcaccgat acgcgagcga acgtgaagcg actgctgctg
     4621 caaaacgtct gcgacctgag caacaacatg aatggtcttc ggtttccgtg tttcgtaaag
     4681 tctggaaacg cggaagtcag cgccctgcac cattatgttc cggatctgca tcgcaggatg
     4741 ctgctggcta ccctgtggaa cacctacatc tgtattaacg aagcgctggc attgaccctg
     4801 agtgattttt ctctggtccc gccgcatcca taccgccagt tgtttaccct cacaacgttc
     4861 cagtaaccgg gcatgttcat catcagtaac ccgtatcgtg agcatcctct ctcgtttcat
     4921 cggtatcatt acccccatga acagaaatcc cccttacacg gaggcatcag tgaccaaaca
     4981 ggaaaaaacc gcccttaaca tggcccgctt tatcagaagc cagacattaa cgcttctgga
     5041 gaaactcaac gagctggacg cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca
     5101 cgctgatgag ctttaccgca gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg
     5161 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     5221 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc
     5281 acgtagcgat agcggagtgt atactggctt aactatgcgg catcagagca gattgtactg
     5341 agagtgcacc attgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     5401 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5461 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5521 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5581 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5641 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5701 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     5761 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     5821 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     5881 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     5941 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6001 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     6061 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     6121 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     6181 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     6241 ttttggtcat gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg
     6301 agccatattc aacgggaaac gtcttgctct aggccgcgat taaattccaa catggatgct
     6361 gatttatatg ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat
     6421 cgattgtatg ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt
     6481 gccaatgatg ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt
     6541 ccgaccatca agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc
     6601 cccgggaaaa cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt
     6661 gatgcgctgg cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt
     6721 aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt
     6781 gatgcgagtg attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa
     6841 atgcataaac ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt
     6901 gataacctta tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga
     6961 atcgcagacc gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct
     7021 tcattacaga aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg
     7081 cagtttcatt tgatgctcga tgagtttttc taagaattaa ttcatgagcg gatacatatt
     7141 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     7201 acctaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     7261 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     7321 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     7381 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     7441 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     7501 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     7561 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     7621 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc a
//
