LOCUS       Exported                5449 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Encodes C-terminal part of RANKL (aa158-aa316) with a GST N-terminal
            tag.
ACCESSION   .
VERSION     .
KEYWORDS    pGEX4T2 RANK-L
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5449)
  AUTHORS   Beranger GE, Momier D, Guigonis JM, Samson M, Carle GF, Scimeca JC
  TITLE     Differential binding of poly(ADP-Ribose) polymerase-1 and JunD/Fra2 
            accounts for RANKL-induced Tcirg1 gene expression during 
            osteoclastogenesis.
  JOURNAL   J Bone Miner Res. 2007 Jul;22(7):975-83. doi: 10.1359/jbmr.070406.
  PUBMED    17419679
REFERENCE   2  (bases 1 to 5449)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5449
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        183..211
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1499..1521)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1666..1684)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1752..1856
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1857..2717
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2075..2094)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2888..3476
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3377..3396
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3630..3647
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3720..3797
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             3798..4880
                     /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(3817..3836)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4893..4914
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4929..4959
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4967..4983
                     /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     4991..5007
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4991..5007
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             5003..81
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5023..5040)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5023..5039)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5032..5054)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatccccagg aattcccggg tcgacaaagc
      961 ctgaggccca gccatttgca cacctcacca tcaatgctgc cagcatccca tcgggttccc
     1021 ataaagtcac tctgtcctct tggtaccacg atcgaggctg ggccaagatc tctaacatga
     1081 cgttaagcaa cggaaaacta agggttaacc aagatggctt ctattacctg tacgccaaca
     1141 tttgctttcg gcatcatgaa acatcgggaa gcgtacctac agactatctt cagctgatgg
     1201 tgtatgtcgt taaaaccagc atcaaaatcc caagttctca taacctgatg aaaggaggga
     1261 gcacgaaaaa ctggtcgggc aattctgaat tccactttta ttccataaat gttgggggat
     1321 ttttcaagct ccgagctggt gaagaaatta gcattcaggt gtccaaccct tccctgctgg
     1381 atccggatca agatgcgacg tactttgggg ctttcaaagt tcaggacata gactgagcgg
     1441 ccgcgcatcg tgactgactg acgatctgcc tcgcgcgttt cggtgatgac ggtgaaaacc
     1501 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     1561 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc
     1621 agtcacgtag cgatagcgga gtgtataatt cttgaagacg aaagggcctc gtgatacgcc
     1681 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     1741 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     1801 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     1861 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     1921 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     1981 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     2041 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
     2101 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     2161 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     2221 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     2281 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     2341 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     2401 cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     2461 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     2521 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     2581 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     2641 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     2701 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     2761 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     2821 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2881 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2941 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     3001 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     3061 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     3121 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     3181 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     3241 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     3301 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     3361 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     3421 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     3481 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     3541 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     3601 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     3661 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc ataaattccg
     3721 acaccatcga atggtgcaaa acctttcgcg gtatggcatg atagcgcccg gaagagagtc
     3781 aattcagggt ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag tatgccggtg
     3841 tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct gcgaaaacgc
     3901 gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc gtggcacaac
     3961 aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg gccctgcacg
     4021 cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg
     4081 tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc
     4141 tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag gatgccattg
     4201 ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct gaccagacac
     4261 ccatcaacag tattattttc tcccatgaag acggtacgcg actgggcgtg gagcatctgg
     4321 tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct gtctcggcgc
     4381 gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg
     4441 aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg
     4501 agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc
     4561 gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg
     4621 ataccgaaga cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc
     4681 tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg
     4741 gcaatcagct gttgcccgtc tcactggtga aaagaaaaac caccctggcg cccaatacgc
     4801 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     4861 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca
     4921 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     4981 caatttcaca caggaaacag ctatgaccat gattacggat tcactggccg tcgttttaca
     5041 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     5101 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     5161 cagcctgaat ggcgaatggc gctttgcctg gtttccggca ccagaagcgg tgccggaaag
     5221 ctggctggag tgcgatcttc ctgaggccga tactgtcgtc gtcccctcaa actggcagat
     5281 gcacggttac gatgcgccca tctacaccaa cgtaacctat cccattacgg tcaatccgcc
     5341 gtttgttccc acggagaatc cgacgggttg ttactcgctc acatttaatg ttgatgaaag
     5401 ctggctacag gaaggccaga cgcgaattat ttttgatggc gttggaatt
//
