Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V012124 | pBIG1a | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pBIG1a
- Antibiotic Resistance:
- Ampicillin, Spectinomycin
- Length:
- 5915 bp
- Type:
- Insect Expression
- Replication origin:
- ori
- Selection Marker:
- Gentamicin
- Promoter:
- Pc
- Cloning Method:
- Gibson Cloning
- 5' Primer:
- TCAACAGGTTGAACTGCTGATC
- 3' Primer:
- GGTGTAGCGTCGTAAGCTAATAC
pBIG1a vector Map
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5. Store the plasmid at -20 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pBIG1a vector Sequence
LOCUS 40924_6502 5915 bp DNA circular SYN 13-JUL-2021
DEFINITION pBIG1a - Step1 vector a (biGBac system for expression of protein
complexes in insect cells).
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5915)
AUTHORS Weissmann F, Petzold G, VanderLinden R, Huis In 't Veld PJ, Brown
NG, Lampert F, Westermann S, Stark H, Schulman BA, Peters JM
TITLE biGBac enables rapid gene assembly for the expression of large
multisubunit protein complexes.
JOURNAL Proc Natl Acad Sci U S A. 2016 May 10;113(19):E2564-9. doi:
10.1073/pnas.1604935113. Epub 2016 Apr 25.
PUBMED 27114506
REFERENCE 2 (bases 1 to 5915)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 5915)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1073/pnas.1604935113"; journalName: "Proc Natl Acad Sci U S A";
date: "2016-05-10- 10"; volume: "113"; issue: "19"; pages: "E2564-9"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..5915
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS 442..1230
/codon_start=1
/label=SmR
/note="aminoglycoside adenylyltransferase (Murphy, 1985)"
/translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
rep_origin 1274..1728
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 1754..1858
/label=AmpR promoter
CDS 1859..2716
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 2890..3478
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3632..3649
/label=L4440
/note="L4440 vector, forward primer"
mobile_element complement(3783..4007)
/label=Tn7R
/note="mini-Tn7 element (right end of the Tn7 transposon)"
CDS complement(4077..4607)
/codon_start=1
/label=GmR
/note="gentamycin acetyltransferase"
/translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
EEVMHFDIDPSTAT"
promoter complement(4796..4824)
/label=Pc promoter
/note="class 1 integron promoter"
protein_bind complement(5220..5253)
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
primer_bind complement(5299..5318)
/label=TK-pA-R
/note="Thymidine kinase polyA, reverse primer"
polyA_signal 5343..5391
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
mobile_element complement(5668..5833)
/label=Tn7L
/note="mini-Tn7 element (left end of the Tn7 transposon)"