LOCUS       sequence-93127-e        4754 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for solubility screening of proteins or protein 
            fragments using a split-GFP assay..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4754)
  AUTHORS   Bulloch EM, Kingston RL
  TITLE     Identifying protein domains by global analysis of soluble fragment 
            data.
  JOURNAL   Anal Biochem. 2014 Jul 10. pii: S0003-2697(14)00280-2. doi: 
            10.1016/j.ab.2014.06.021.
  PUBMED    25016192
REFERENCE   2  (bases 1 to 4754)
  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: "Anal 
            Biochem. 2014 Jul 10. pii: S0003-2697(14)00280-2. doi: 
            10.1016/j.ab.2014.06.021."
COMMENT     Sequence Label: pBAD_GFP11_T7LysH17A
FEATURES             Location/Qualifiers
     source          1..4754
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             27..905
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     CDS             1042..1497
                     /codon_start=1
                     /product="lysozyme from bacteriophage T7"
                     /label=T7 lysozyme
                     /note="inhibitor of T7 RNA polymerase"
                     /translation="MARVQFKQRESTDAIFVACSATKPSQNVGVREIRQWHKEQGWLDV
                     GYHFIIKRDGTVEAGRDEMAVGSHAKGYNHNSIGVCLVGGIDDKGKFDANFTPAQMQSL
                     RSLLVTLLAKYEGAVLRAHHEVAPKACPSFDLKRWWEKNELVTSDRG"
     promoter        1495..1528
                     /label=T7 Phi-3.8 promoter
                     /note="weak promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1703..1725)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1866..1885
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1965..1982)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2136..2724)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2216..2235)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2895..3686,3687..3755))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3518..3537
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3756..3847)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(3867..3894)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3986..4072)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     4225..4242
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(4302..4349)
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     CDS             complement(4407..4424)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        complement(4470..4754)
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     complement(4528..4547)
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 ctctgaatgg cgggagtatg aaaagtatgg ctgaagcgca aaatgatccc ctgctgccgg
       61 gatactcgtt taatgcccat ctggtggcgg gtttaacgcc gattgaggcc aacggttatc
      121 tcgatttttt tatcgaccga ccgctgggaa tgaaaggtta tattctcaat ctcaccattc
      181 gcggtcaggg ggtggtgaaa aatcagggac gagaatttgt ttgccgaccg ggtgatattt
      241 tgctgttccc gccaggagag attcatcact acggtcgtca tccggaggct cgcgaatggt
      301 atcaccagtg ggtttacttt cgtccgcgcg cctactggca tgaatggctt aactggccgt
      361 caatatttgc caatacgggg ttctttcgcc cggatgaagc gcaccagccg catttcagcg
      421 acctgtttgg gcaaatcatt aacgccgggc aaggggaagg gcgctattcg gagctgctgg
      481 cgataaatct gcttgagcaa ttgttactgc ggcgcatgga agcgattaac gagtcgctcc
      541 atccaccgat ggataatcgg gtacgcgagg cttgtcagta catcagcgat cacctggcag
      601 acagcaattt tgatatcgcc agcgtcgcac agcatgtttg cttgtcgccg tcgcgtctgt
      661 cacatctttt ccgccagcag ttagggatta gcgtcttaag ctggcgcgag gaccaacgta
      721 tcagccaggc gaagctgctt ttgagcacca cccggatgcc tatcgccacc gtcggtcgca
      781 atgttggttt tgacgatcaa ctctatttct cgcgggtatt taaaaaatgc accggggcca
      841 gcccgagcga gttccgtgcc ggttgtgaag aaaaagtgaa tgatgtagcc gtcaagttgt
      901 cataattggt aacgaatcag acaattgacg gcttgacgga gtagcatagg gtttgcagaa
      961 tccctgcttc gtccatttga caggcacatt atgcatgcgg gtcccctttg atagattaaa
     1021 aaggaaagga ggaaagaaat aatggctcgt gtacagttta aacaacgtga atctactgac
     1081 gcaatctttg ttgcctgctc ggctaccaag ccaagtcaga atgttggtgt ccgtgagatt
     1141 cgccagtggc acaaagagca gggttggctc gatgtgggat accactttat catcaagcga
     1201 gacggtactg tggaggcagg acgagatgag atggctgtag gctctcacgc taagggttac
     1261 aaccacaact ctatcggcgt ctgccttgtt ggtggtatcg acgataaagg taagttcgac
     1321 gctaacttta cgccagccca aatgcaatcc cttcgctcac tgcttgtcac actgctggct
     1381 aagtacgaag gcgctgtgct tcgcgcccat catgaggtgg cgccgaaggc ttgcccttcg
     1441 ttcgacctta agcgttggtg ggagaagaac gaactggtca cttctgaccg tggataatta
     1501 attgaactca ctaaagggag accacagcgg tttccctttg ttcgcattgg aggtcaaata
     1561 atgcgcaagt cttataaaca attctataag gctccgagga ggcatatcca agtgtgggag
     1621 gcagccaatg ggccgcatgc cgcttcgcct tcgcgcgcga attgatctgc tgcctcgcgc
     1681 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     1741 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     1801 ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa
     1861 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
     1921 cagatgcgta aggagaaaat accgcatcag cgctcttccg cttcctcgct cactgactcg
     1981 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     2041 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     2101 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     2161 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     2221 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     2281 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     2341 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     2401 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     2461 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     2521 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     2581 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     2641 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     2701 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     2761 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     2821 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     2881 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     2941 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     3001 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     3061 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     3121 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     3181 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3241 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3301 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     3361 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     3421 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     3481 cggcgaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc acatagcaga
     3541 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     3601 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     3661 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     3721 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     3781 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     3841 aaacaaaaag agtttgtaga aacgcaaaaa ggccatccgt caggatggcc ttctgcttaa
     3901 tttgatgcct ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc cgttgcttcg
     3961 caacgttcaa atccgctccc ggcggatttg tcctactcag gagagcgttc accgacaaac
     4021 aacagataaa acgaaaggcc cagtctttcg actgagcctt tcgttttatt tgatgcctgg
     4081 cagttcccta ctctcgcatg gggagacccc acactaccat cggcgctacg gcgtttcact
     4141 tctgagttcg gcatggggtc aggtgggacc accgcgctac tgccgccagg caaattctgt
     4201 tttatcagac cgcttctgcg ttctgattta atctgtatca ggctgaaaat cttctctcat
     4261 ccgccaaaac agccaagctc acgagtgcgg ccgcaagctt atgtgatgcc agcagcattt
     4321 acgtactcat gaagaaccat atggtcacga cttgttgaac cagaaccaga gcctccatcc
     4381 tcgagcagct gactagtgct accaccatga tgatgatgat gatgagaacc gcccatggtt
     4441 aattcctcct gttagcccaa aaaacgggta tggagaaaca gtagagagtt gcgataaaaa
     4501 gcgtcaggta ggatccgcta atcttatgga taaaaatgct atggcatagc aaagtgtgac
     4561 gccgtgcaaa taatcaatgt ggacttttct gccgtgatta tagacacttt tgttacgcgt
     4621 ttttgtcatg gctttggtcc cgctttgtta cagaatgctt ttaataagcg gggttaccgg
     4681 tttggttagc gagaagagcc agtaaaagac gcagtgacgg caatgtctga tgcaatatgg
     4741 acaattggtt tctt
//
