LOCUS       sequence-358130-        5098 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor with homologous arms for Rab11a to knock in SFFV-T2A-Puromycin
            resistance-PolyA cassette (for knock out.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5098)
  AUTHORS   Gemperle J, Harrison TS, Flett C, Adamson AD, Caswell PT
  TITLE     On demand expression control of endogenous genes with DExCon, 
            DExogron and LUXon reveals differential dynamics of Rab11 family 
            members.
  JOURNAL   Elife. 2022 Jun 16;11. pii: 76651. doi: 10.7554/eLife.76651.
  PUBMED    35708998
REFERENCE   2  (bases 1 to 5098)
  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: "Elife."; 
            date: "2022-06-16"; volume: "11. pii: 76651. doi"; pages: " 
            10.7554/eLife.76651"
COMMENT     Sequence Label: pJET-61-SFFV-T2A-PuroR-polyA-R11a
FEATURES             Location/Qualifiers
     source          1..5098
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     111..130
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     364..381
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    501..522
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(537..542,543..560,561..567)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     promoter        1008..1026
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        1353..1760
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     CDS             1859..1912
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     complement(2553..2570)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2559..2783
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(3174..3192)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3498..3516)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3584..3688
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3689..3757,3758..4549)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3907..3926)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(4720..5098,1..210)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
       61 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
      121 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
      181 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
      241 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
      301 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
      361 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
      421 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcaatt
      481 ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta
      541 cactttatgc ttccggctcg tataatgtgt ggaattatga gcggataata atttcacaca
      601 ggaggtttaa actttaaaca tgtcaaaaga gacgtctttt gttaagaatg ctgaggaact
      661 tgcaaagcaa aaaatggatg ctattaaccc tgaactttct tcaaaattta aatttttaat
      721 aaaattcctg tctcagtttc ctgaagcttg ctctaaacct cgttcaaaaa aaatgcagaa
      781 taaagttggt caagaggaac atattgaata tttagctcgt agttttcatg agagtcgatt
      841 gccaagaaaa cccacgccac ctacaacggt tcctgatgag gtggttagca tagttcttaa
      901 tataagtttt aatatacagc ctgaaaatct tgagagaata aaagaagaac atcgattttc
      961 catggcagct gagaatattg taggagatct tctagaaaga tagctgataa tacgactcac
     1021 tatagggact atggtagcta ggagttccag gactcagttt cccctttgag cctcctttag
     1081 cgactaaagc ttgaagcccc acgcatctcg actctcgcgc acaccgccct tgttgggctc
     1141 aggggcgggg cgccgccccc ggaagtactt ccccttaaag gctggggcct gccggaaatg
     1201 gcgcagcggc agggaggggc tcttcaccca gtccggcagt tgaagctcgg cgctcgggtt
     1261 acccctgcag cgacgccccc tggtcccaca gataccactg ctgctcccgc cctttcgctc
     1321 ctcggccgcg caatgggatc gggtgggact aggtaacgcc attttgcaag gcatggaaaa
     1381 ataccaaacc aagaatagag aagttcagat caagggcggg tacatgaaaa tagctaacgt
     1441 tgggccaaac aggatatctg cggtgagcag tttcggcccc ggcccggggc caagaacaga
     1501 tggtcaccgc agtttcggcc ccggcccgag gccaagaaca gatggtcccc agatatggcc
     1561 caaccctcag cagtttctta agacccatca gatgtttcca ggctccccca aggacctgaa
     1621 atgaccctgc gccttatttg aattaaccaa tcagcctgct tctcgcttct gttcgcgcgc
     1681 ttctgcttcc cgagctctat aaaagagctc acaacccctc actcggcgcg ccagtcctcc
     1741 gacagactga gtcgcccggg ggggatctgg agctctcgag aattctcacg cgtctgcagg
     1801 atatcaagct tgcggtaccg cgggcccggg atccatggtg agcactagtg gcagcggaga
     1861 gggcagagga agtcttctaa catgcggtga cgtggaggag aatcccggcc ctaggctcga
     1921 gatgacggaa tataagccca cagtcagatt ggctacgaga gatgatgttc cgcgagccgt
     1981 gcggacgctc gcagccgctt ttgcagacta tcctgctacg cgacatactg ttgaccctga
     2041 taggcatatc gaaagagtaa ccgaacttca ggagctcttt ctgactaggg tcggtttgga
     2101 tattgggaag gtctgggtag ccgacgacgg cgccgctgtc gccgtatgga cgactccaga
     2161 gagcgtagaa gcgggagcgg tatttgctga gatcggtcca cggatggcag aattgtccgg
     2221 cagtagactt gcggctcaac agcaaatgga aggacttctc gcaccgcatc gacccaaaga
     2281 gcccgcctgg tttcttgcta cggtaggggt gtcccccgac caccagggta agggtctcgg
     2341 aagtgctgtt gtccttccag gagtggaagc ggcagagcgc gcgggtgttc ctgcgtttct
     2401 ggagacctca gccccaagga accttccttt ttatgaacgc ttgggattta ctgtcacggc
     2461 cgatgttgag gttcctgaag gacctcggac atggtgcatg acaaggaaac caggggcgtg
     2521 atctagaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     2581 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     2641 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     2701 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     2761 tggggatgcg gtgggctcta tgggtcgacg gaaccggtgc tggaagtggt acacgcgacg
     2821 acgagtacga ctacctcttt aaaggtgagg ccatgggctc tcgcactcta cacagtcctc
     2881 gttcggggac ccgggccact cccggtggac cctcgtgccg gccacccctg cactgatata
     2941 ggcctccctc agcccttcct ttttgtgcgg ttccgtctcc tacccagctc agcctcttct
     3001 cccccgctca gacaggggtc cccatcacat gccgctctct gagcgacctc tccataggcc
     3061 ttcgctggcc tcagagcccc tccctgcgtg tccttcccct ggcggactgc cttctcccac
     3121 atcgtatctt gctgaaaaac tcgagccatc cggaagatct ggcggccgct ctccctatag
     3181 tgagtcgtat tacgccggat ggatatggtg ttcaggcaca agtgttaaag cagttgattt
     3241 tattcactat gatgaaaaaa acaatgaatg gaacctgctc caagttaaaa atagagataa
     3301 taccgaaaac tcatcgagta gtaagattag agataataca acaataaaaa aatggtttag
     3361 aacttactca cagcgtgatg ctactaattg ggacaatttt ccagatgaag tatcatctaa
     3421 gaatttaaat gaagaagact tcagagcttt tgttaaaaat tatttggcaa aaataatata
     3481 attcggctgc aggggcggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     3541 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     3601 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     3661 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     3721 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     3781 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     3841 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     3901 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     3961 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     4021 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     4081 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     4141 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     4201 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     4261 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     4321 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     4381 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     4441 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     4501 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     4561 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     4621 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     4681 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     4741 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     4801 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     4861 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     4921 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     4981 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5041 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatac
//
