LOCUS       sequence-482348-        6643 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Expresses GRP78 with the last 73 amino acids at the C terminus 
            deleted fused to the GST Tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6643)
  AUTHORS   Tseng CC, Zhang P, Lee AS
  TITLE     The COOH-Terminal Proline-Rich Region of GRP78 Is a Key Regulator of
            Its Cell Surface Expression and Viability of Tamoxifen-Resistant 
            Breast Cancer Cells.
  JOURNAL   Neoplasia. 2019 Aug;21(8):837-848. doi: 10.1016/j.neo.2019.05.008. 
            Epub 2019 Jul 12.
  PUBMED    31306849
REFERENCE   2  (bases 1 to 6643)
  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"; doi: 
            "10.1016/j.neo.2019.05.008"; journalName: "Neoplasia"; date: 
            "2019-08"; volume: "21"; issue: "8"; pages: "837-848"
COMMENT     Sequence Label: GST-GRP78-Delta C73 (pGEX4T1)
FEATURES             Location/Qualifiers
     source          1..6643
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(181..186,187..202,203..209)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    217..233
                     /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             256..909
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     867..889
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             916..933
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2691..2713)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2858..2876)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2944..3048
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3049..3117,3118..3909)
                     /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(3267..3286)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4080..4668
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4569..4588
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4822..4839
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4912..4989
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4990..6072
                     /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(5009..5028)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6121..6126,6127..6144,6145..6151)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6159..6175
                     /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     6183..6199
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6183..6199
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(6195..6643,1..79)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6215..6232)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6215..6231)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6224..6246)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 cttatcgact gcacggtgca ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt
       61 atggctgtgc aggtcgtaaa tcactgcata attcgtgtcg ctcaaggcgc actcccgttc
      121 tggataatgt tttttgcgcc gacatcataa cggttctggc aaatattctg aaatgagctg
      181 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
      241 caggaaacag tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
      301 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
      361 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
      421 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
      481 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
      541 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
      601 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
      661 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
      721 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
      781 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
      841 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
      901 cctccaaaat cggatctggt tccgcgtgga tccgaggagg aggacaagaa ggaggacgtg
      961 ggcacggtgg tcggcatcga cctggggacc acctactcct gcgtcggcgt gttcaagaac
     1021 ggccgcgtgg agatcatcgc caacgatcag ggcaaccgca tcacgccgtc ctatgtcgcc
     1081 ttcactcctg aaggggaacg tctgattggc gatgccgcca agaaccagct cacctccaac
     1141 cccgagaaca cggtctttga cgccaagcgg ctcatcggcc gcacgtggaa tgacccgtct
     1201 gtgcagcagg acatcaagtt cttgccgttc aaggtggttg aaaagaaaac taaaccatac
     1261 attcaagttg atattggagg tgggcaaaca aagacatttg ctcctgaaga aatttctgcc
     1321 atggttctca ctaaaatgaa agaaaccgct gaggcttatt tgggaaagaa ggttacccat
     1381 gcagttgtta ctgtaccagc ctattttaat gatgcccaac gccaagcaac caaagacgct
     1441 ggaactattg ctggcctaaa tgttatgagg atcatcaacg agcctacggc agctgctatt
     1501 gcttatggcc tggataagag ggagggggag aagaacatcc tggtgtttga cctgggtggc
     1561 ggaaccttcg atgtgtctct tctcaccatt gacaatggtg tcttcgaagt tgtggccact
     1621 aatggagata ctcatctggg tggagaagac tttgaccagc gtgtcatgga acacttcatc
     1681 aaactgtaca aaaagaagac gggcaaagat gtcaggaaag acaatagagc tgtgcagaaa
     1741 ctccggcgcg aggtagaaaa ggccaaacgg gccctgtctt ctcagcatca agcaagaatt
     1801 gaaattgagt ccttctatga aggagaagac ttttctgaga ccctgactcg ggccaaattt
     1861 gaagagctca acatggatct gttccggtct actatgaagc ccgtccagaa agtgttggaa
     1921 gattctgatt tgaagaagtc tgatattgat gaaattgttc ttgttggtgg ctcgactcga
     1981 attccaaaga ttcagcaact ggttaaagag ttcttcaatg gcaaggaacc atcccgtggc
     2041 ataaacccag atgaagctgt agcgtatggt gctgctgtcc aggctggtgt gctctctggt
     2101 gatcaagata caggtgacct ggtactgctt gatgtatgtc cccttacact tggtattgaa
     2161 actgtgggag gtgtcatgac caaactgatt ccaaggaaca cagtggtgcc taccaagaag
     2221 tctcagatct tttctacagc ttctgataat caaccaactg ttacaatcaa ggtctatgaa
     2281 ggtgaaagac ccctgacaaa agacaatcat cttctgggta catttgatct gactggaatt
     2341 cctcctgctc ctcgtggggt cccacagatt gaagtcacct ttgagataga tgtgaatggt
     2401 attcttcgag tgacagctga agacaagggt acagggaaca aaaataagat cacaatcacc
     2461 aatgaccaga atcgcctgac acctgaagaa atcgaaagga tggttaatga tgctgagaag
     2521 tttgctgagg aagacaaaaa gctcaaggag cgcattgata ctagaaatga gttggaaagc
     2581 tatgcctatt ctctaaagaa tcagattgga gataaagaaa agtagctcga gcggccgcat
     2641 cgtgactgac tgacgatctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     2701 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     2761 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt
     2821 agcgatagcg gagtgtataa ttcttgaaga cgaaagggcc tcgtgatacg cctattttta
     2881 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     2941 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     3001 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     3061 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     3121 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     3181 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     3241 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tgttgacgcc
     3301 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     3361 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     3421 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     3481 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     3541 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg
     3601 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     3661 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     3721 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     3781 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     3841 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     3901 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     3961 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4021 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4081 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4141 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4201 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4261 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4321 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4381 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4441 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4501 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4561 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4621 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4681 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     4741 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     4801 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     4861 cggtattttc tccttacgca tctgtgcggt atttcacacc gcataaattc cgacaccatc
     4921 gaatggcgca aaacctttcg cggtatggca tgatagcgcc cggaagagag tcaattcagg
     4981 gtggtgaatg tgaaaccagt aacgttatac gatgtcgcag agtatgccgg tgtctcttat
     5041 cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac gcgggaaaaa
     5101 gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca acaactggcg
     5161 ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca cgcgccgtcg
     5221 caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt ggtggtgtcg
     5281 atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct tctcgcgcaa
     5341 cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat tgctgtggaa
     5401 gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac acccatcaac
     5461 agtattattt tctcccatga agacggtacg cgactgggcg tggagcatct ggtcgcattg
     5521 ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc gcgtctgcgt
     5581 ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa
     5641 ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc
     5701 gttcccactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt
     5761 accgagtccg ggctgcgcgt tggtgcggat atctcggtag tgggatacga cgataccgaa
     5821 gacagctcat gttatatccc gccgttaacc accatcaaac aggattttcg cctgctgggg
     5881 caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa gggcaatcag
     5941 ctgttgcccg tctcactggt gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc
     6001 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
     6061 agcgggcagt gagcgcaacg caattaatgt aagttagctc actcattagg caccccaggc
     6121 tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca
     6181 cacaggaaac agctatgacc atgattacgg attcactggc cgtcgtttta caacgtcgtg
     6241 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
     6301 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga
     6361 atggcgaatg gcgctttgcc tggtttccgg caccagaagc ggtgccggaa agctggctgg
     6421 agtgcgatct tcctgaggcc gatactgtcg tcgtcccctc aaactggcag atgcacggtt
     6481 acgatgcgcc catctacacc aacgtgacct atcccattac ggtcaatccg ccgtttgttc
     6541 ccacggagaa tccgacgggt tgttactcgc tcacatttaa tgttgatgaa agctggctac
     6601 aggaaggcca gacgcgaatt atttttgatg gcgttggaat tag
//
