Bac-to-Bac™ C-His TOPO™ Cloning Kit
본 제품은 LMO 제품으로, 고객 분께서 LMO 신고 시스템을 통해 직접 수입 신고를 진행해주셔야 합니다. 자세히보기
Bac-to-Bac™ C-His TOPO™ Cloning Kit
Gibco™

Bac-to-Bac™ C-His TOPO™ Cloning Kit

Bac-to-Bac™ Baculovirus Expression is an efficient method for producing baculovirus for high-level protein expression in insect cells. It relies on자세히 알아보기
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카탈로그 번호수량
A1109820 Reactions
카탈로그 번호 A11098
제품 가격(KRW)
2,210,000
온라인 행사
Ends: 31-Dec-2025
2,599,000
할인액 389,000 (15%)
Each
카트에 추가하기
수량:
20 Reactions
제품 가격(KRW)
2,210,000
온라인 행사
Ends: 31-Dec-2025
2,599,000
할인액 389,000 (15%)
Each
카트에 추가하기
Bac-to-Bac™ Baculovirus Expression is an efficient method for producing baculovirus for high-level protein expression in insect cells. It relies on generation of recombinant virus by site-specific transposition in E. coli rather than homologous recombination in insect cells. New: pFastBacTOPO™ vectors!


Reliable and fast protein expression
The Bac-to-Bac™ Baculovirus Expression System is faster and easier than traditional baculovirus expression, and it maintains high levels of protein expression. Bac-to-Bac™ relies on generation of recombinant baculovirus by site-specific transposition in E. coli rather than homologous recombination in insect cells. The Bac-to-Bac™ Baculvirus Expression System is highly regarded in academic Literature. Approximately 100 citations every year since 2000!

Traditional baculovirus systems require purification and amplification of an initial low-titer viral supernatant. This requires a time-consuming plaque assay. Bac-to-Bac™'s pFastBac™ vector, however, recombines with the parent bacmid in DH10Bac™ E. coli competent cells to form an expression bacmid. Transfect the bacmid into insect cells for fast production of a high titer of pure recombinant baculovirus particles in the very first transfection. You'll save weeks of precious time. Collect a pure P2 baculovirus stock on Day 10 without the necessity of tedious plaques assay (See Figure 1)

High expression, easy screening
pFastBac™ uses the strong polyhedrin promoter to generate high levels of expression in a variety of insect cell lines such as Sf9, Sf21, and High Five™ cells.

New additions:
Bac-to-Bac™'s pFastBac vectors are now available as pFastBacTOPO™ vectors too. pFastBacTOPO™ vectors are Blunt™ TOPO™ vectors and are supplied with Mach1™-T1R E. coli for easy cloning and pFastBac⁄GOI propagation. The new vectors enable you to:

• Amplify your gene of interest (GOI) with a PCR enzyme of highest fidelity such as Accuprime™ Pfx SuperMix.

• TOPO™ clone the blunt-end PCR product into the new vector in only 5 minutes!

• Visualize colonies 8 hours after plating on ampicillin selective plates because Mach1™-T1R E. coli cells have a faster doubling time compared to other standard cloning strains.

• With the Bac-to-Bac™ C-His TOPO™ cloning or expression kit, you can produce a C-terminal His-fusion protein with a TEV cleavage site to purify with nickel-chelating resins (Invitrogen’s ProBond™ Purification System) and generate a native protein with the aid of Invitrogen’s popular AcTEV™ Protease. This is the Invitrogen’s only vector that contains a C-terminal TEV cleavage site!

The new vectors combine the TOPO™ cloning technology with the highly regarded Bac-to-Bac™ baculovirus expression features to enable easy cloning and high-level protein expression.
The pFastBacTOPO™ vectors are offered as cloning and expression kits.
For Research Use Only. Not for use in diagnostic procedures.
사양
항생제 내성 박테리아Ampicillin (AmpR), Gentamicin (GmR)
세포주High Five™, Sf21, Sf9
세포 유형Insect Cell
클로닝 방법Blunt TOPO
발현 메커니즘Insect Cell Expression
발현 시스템Baculovirus
용도(애플리케이션)Protein Expression, Viral Vaccine Production
반응 수20 Reactions
제품라인Bac-to-Bac, TOPO
제품 유형Cloning Kit
프로모터Polyhedrin
단백질 태그His Tag (6x)
수량20 Reactions
선택 제제(진핵)Ampicillin, Gentamicin
벡터pFastBac/CT-TOPO
Unit SizeEach
구성 및 보관
Kit content for Bac-to-Bac™ C-His TOPO™ cloning kit (For details see the manual)
• Vector kit for 20 topo cloning reactions:
o pFastBac⁄CT-TOPO™ Vector containing the C-terminal TEV cleavage sit and His-Tag.
o pFastBac⁄CT-Gus Control Plasmid (Control expression vector)
o Other reagents supplied: 10x PCR, dNTP Mix, Salt Solution, Sterile Water, Control PCR template, Polyhedrin Forward Primer, SV40 pA Reverse Primer
• Kit containing competent cells (20 reactions):
o One Shot™ Mach1-T1R Chemically Competent E. coli

Vector kit: store at -20°C
One Shot™ Mach1-T1R Chemically Competent E. coli: store at -80°C

자주 묻는 질문(FAQ)

I cannot detect any recombinant fusion protein after using the BaculoDirect Expression Kit. What could be the cause for this and what do you suggest I try?

Please check the construction of your entry clone, and ensure that the insert is in frame with the vector. Analyze the recombinant viral DNA by PCR to confirm the correct size and orientation of your insert after the LR reaction. Sequence your PCR product to verify the proper reading frame for expression of the epitope tag.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

I'm getting a low-titer P1 viral stock and would like to generate a high-titer stock. What should I do?

To get a high-titer stock, reinfect cells with the P1 stock and generate a P2 high-titer stock. Follow the directions in the BaculoDirect manual on page 18 to generate your P2 stock.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

I performed an LR reaction, followed by transfection into Sf9/Sf21 insect cells, but do not see any signs of infection even though it's been 72 hours. What should i do?

Please see our recommendations below:

- Check the LR reaction by PCR analysis prior to transfection into insect cells.
- We recommend using Grace's Insect Cell Culture Medium, Unsupplemented during the transfection experiment instead of serum-free medium, as components in serum-free medium may interfere with transfection.
- Ensure that FBS, supplements, or antibiotics are not included during transfection, as the proteins in these materials can interfere with the Cellfectin II Reagent.
- Use the LR recombination reaction using the pENTR/CAT plasmid as a positive control and Cellfectin II Reagent only (mock transfection) as a negative control.
- Ensure that cells are in the log phase of growth with >95% viability, and the amount of cells are in accordance with the suggestions in the manual.
- Cells may not show signs of viral infection for up to a week depending on transfection efficiency; continue culturing and monitor cells daily for signs of infection.

I see a precipitate in my ganciclovir solution. What can I do?

Warm the ganciclovir solution in a water bath at 37 degrees C for 5-10 min, then vortex for a few minutes. The precipitate should go back into solution.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

I am purifying my secreted protein expressed in insect cells with a His-tagged purification column and getting no yield of my protein. Why and what can I do?

Media used to culture insect cells usually have an acidic pH (6.0-6.5) or contain electron-donating groups that can prevent binding of the 6xHis-tagged protein to Ni-NTA. Amino acids such as glutamine, glycine, or histidine are present at significantly higher concentrations in media for growing insect cells than in media for growing mammalian cells, and compete with the 6xHis-tagged protein for binding sites on Ni-NTA matrices. Grace's medium (Thermo Fisher Scientific), for example, contains approximately 10 mM glutamine, 10 mM glycine, and 15 mM histidine.

Dialysis of the medium against a buffer with the appropriate composition and pH (8.0) similar to the lysis buffer recommended for purification under native conditions usually restores optimal binding conditions. Note that depending on the medium used, a white precipitate (probably made up of insoluble salts) can occur, but normally the 6xHis-tagged protein remains in solution. This can be tested by either protein quantitation if using a protein-free medium or by monitoring the amount of 6xHis-tagged protein by western-blot analysis. After centrifugation, 6xHis-tagged protein can be directly purified from the cleared supernatant.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.