pBlue TOPO™ TA Expression Kit

El kit de expresión pBlue TOPO® TA utiliza la expresión de β-galactosidasa para analizar una amplia gama de actividad transcripcionalMás información

Número de catálogo	Cantidad
K483101 también denominado K4831-01	20 reacciones

Número de catálogo K483101

también denominado K4831-01

Precio (CLP)

Cantidad:

20 reacciones

El kit de expresión pBlue TOPO® TA utiliza la expresión de β-galactosidasa para analizar una amplia gama de actividad transcripcional a partir de fragmentos de promotores. Los ensayos para β-galactosidasa son fáciles de realizar y cuantitativos a niveles muy bajos de expresión. El kit de expresión pBlue TOPO® TA es ideal para el análisis funcional de promotores con baja actividad transcripcional.

Para uso exclusivo en investigación. No apto para uso en procedimientos diagnósticos.

Especificaciones

Sistema constitutivo o inducibleConstitutivo

Tipo de entregaTransfección

Para utilizar con (aplicación)Ensayos con indicador, clone su propio promotor

Tipo de productoKit de expresión TOPO TA

Cantidad20 reacciones

Gen marcadorBeta-gal (lacZ)

Agente de selección (eucariótico)Geneticin™ (G-418)

VectorpBlue, vectores TOPO-TA

Método de clonaciónTOPO™-TA

Línea de productosTOPO

PromotorNinguno (sin promoción)

Etiqueta de proteínaSin etiquetar

Unit Size20 reactions

Contenido y almacenamiento

El kit de expresión pBlue TOPO™ TA contiene dos cajas. La caja de clonación TOPO™ TA contiene un vector pBlue-TOPO™ linealizado y activado con la topoisomerasa I, dNTP, primers para la secuenciación, una solución salina, y una plantilla de PCR de control y primers. Esta caja debe almacenarse a -20 °C. La caja One Shot™ contiene células de E. coli One Shot™ TOP10, medio SOC y un plásmido de control superenrollado; almacenar a -80 °C. Se garantiza la estabilidad de los kits durante 6 meses si se almacenan correctamente.

Preguntas frecuentes

Can I store my competent E. coli in liquid nitrogen?

We do not recommend storing competent E. coli strains in liquid nitrogen as the extreme temperature can be harmful to the cells. Also, the plastic storage vials are not intended to withstand the extreme temperature and may crack or break.

How should I store my competent E. coli?

We recommend storing our competent E. coli strains at -80°C. Storage at warmer temperatures, even for a brief period of time, will significantly decrease transformation efficiency.

Do I need to include a ribosomal binding site (RBS/Shine Dalgarno sequence) or Kozak sequence when I clone my gene of interest?

ATG is often sufficient for efficient translation initiation although it depends upon the gene of interest. The best advice is to keep the native start site found in the cDNA unless one knows that it is not functionally ideal. If concerned about expression, it is advisable to test two constructs, one with the native start site and the other with a Shine Dalgarno sequence/RBS or consensus Kozak sequence (ACCAUGG), as the case may be. In general, all expression vectors that have an N-terminal fusion will already have a RBS or initiation site for translation.

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

What is the best molar ratio of PCR product:vector to use for TOPO TA cloning? Is there an equation to calculate the quantity to use?

We suggest starting with a molar ratio of 1:1 (insert:vector), with a range of 0.5:1 to 2:1. The quantity used in a TOPO cloning reaction is typically 5-10 ng of a 2 kb PCR product.

Equation:

length of insert (bp)/length of vector (bp) x ng of vector = ng of insert needed for 1:1 (insert:vector ratio)

What is the best ratio of insert:vector to use for cloning? Is there an equation to calculate this?

The optimal ratio is 1:1 insert to vector. Optimization can be done using a ratio of 0.5-2 molecules of insert for every molecule of the vector.

Equation:

length of insert (bp)/length of vector (bp) x ng of vector = ng of insert needed for 1:1 insert:vector ratio

View all pBlue TOPO™ TA Expression Kit FAQs

Citations & References (1)

Citations & References

Abstract

Mutation of the matrix metalloproteinase At2-MMP inhibits growth and causes late flowering and early senescence in Arabidopsis.

Authors: Golldack Dortje; Popova Olga V; Dietz Karl-Josef;

Journal:J Biol Chem

PubMed ID:11726650

'This study characterizes the expression and functional significance of the member of the matrix metalloproteinase (MMP) family At2-MMP from Arabidopsis. By transcript analysis, expression of At2-MMP was found in leaves and roots of juvenile Arabidopsis and leaves, roots, and inflorescences of mature flowering plants showing strong increase of transcript abundance ... More