Plates were developed using 1 … The insertion site is flanked by BstZI, EcoRI, and NotI sites. The pGEM®-T Vector cloning region is flanked by recognition sites for the enzyme BstZI. Briefly centrifuge the pGEM ®-T or pGEM -T Easy Vector and Control Insert DNA tubes to collect contents at the bottom of the tube. Please try again or contact Customer Service. The high copy number pGEM®-T and pGEM®-T Easy Vectors contain T7 and SP6 RNA polymerase promoters flanking a multiple cloning region within the alpha-peptide coding region of the enzyme beta-galactosidase. When you select your country, you agree that we can place these functional cookies on your device. Abstract. The pGEM ®-T and pGEM -T Easy Vector Systems have been optimized using a 1:1 molar ratio of the Control Insert DNA to the vectors. If initial experiments with your PCR product are suboptimal, ratio optimization may be necessary. This product is available through the Promega Helix onsite stocking program. To protect your privacy, your account will be locked after 6 failed attempts. Stay notified of Promega events, products and news. Latest generation GoTaq® polymerase—high-performance for your everyday PCR needs. There was an issue with the password reset process. Please try again or contact Customer Service. Enter your username and we'll send a link to reset your password. Molecular Cloning: A Laboratory Manual Third Edition. Please update your browser to Internet Explorer 11 or above. Ratios from 3:1 to 1:3 provide good initial parameters. All Rights Reserved. Revised 4/17 www.promega.com 2. Issai Falcon. Note: You will not be able to access your account until your email is verified. After that, you will need to contact Customer Service to unlock your account. Please try again or contact Customer Service. Diese Seite wurde zuletzt am … Instructions for Use of Product(s) A1360, A1380, A3600, A3610. To protect your privacy, your account has been locked after 6 failed login attempts. However, the length of RNA packaged in the virus-like particles with high efficiency is usually less than 500 bases. Please try again or contact Customer Service. The extracellular matrix (ECM) plays an important role in maintaining tissue homeostasis and poses a significant physical barrier to in vivo cell migration. A password reset email has been sent to the primary email address associated with your account. Please try again or contact Customer Service. Background: Development of resistance to doxorubicin-based chemotherapy limits its curative effect in osteosarcoma. Please try again or contact Customer Service. Please try again or contact Customer Service. A resource designed for scientists just embarking on their career, focusing on fundamental technologies and techniques. Thank you for verifying your email address. Ligation Protocol 1. PCR cloning system for expression in mammalian cells. There was an error processing your request. Vector database is a digital collection of vector backbones assembled from publications and commercially available sources. Thank you for verifying your email address. Please check your network settings and try again. There was an error processing your request. Quick PROTOCOL 1 pGEM®-T and pGEM®-T Easy Vector Systems INSTRUCTIONS FOR USE OF PRODUCTS A1360, A1380, A3600 AND A3610. Main. © 2021 Promega Corporation. There was an issue sending the verification email. Please try again or contact Customer Service. Instructions for Use of Product(s) A1360, A1380, A3600, A3610. We prepared a series of pGEM plasmids (Promega) containing 1, 2, 4, 8, 16, 32, or 64 tandem repeats of the sequence described above. Catalog number selected: After that, you will need to contact Customer Service to unlock your account. You've created a Promega.com account. The incubation period may be extended to increase the number of colonies after transformation. The position of the T is indicated by * in the pGEM®-T Vector Sequence (.txt). These single 3´-T overhangs at the insertion site greatly improve the efficiency of ligation of a PCR product into the plasmids by preventing recircularization of the vector and providing a compatible overhang for PCR products generated by certain thermostable polymerases. These samples were collected from Da Longshu village (a, 45 samples), Bai Shiyan village (b, 28 … You have not verified your email address. You've created a Promega.com account. A verified email address is required to access the full functionality of your Promega.com account. Pod pepper (Capsicum frutescens) is widely planted in China, especially around Wenshan city, Yunnan province, and viral diseases have now also become a major threat to pepper production in Yunnan.During July 2019, 89 pepper leaf samples were collected from three different fields in Wenshan. Our website uses functional cookies that do not collect any personal information or track your browsing activity. Please contact Customer Service to unlock your account. Due to our annual Physical inventory orders received after 5PM Central Time on Tuesday March 9 will be shipped according to the following schedule: The green and blue arrows indicate 5′-RACE products characterized from WT and 1–910 EagI constructs. The pGEM®-T Vector is ready to use in ligation reactions, prepared by cutting the pGEM®-5Zf(+) Vector with EcoRV and adding a 3´ terminal thymidine to both ends. PCR cloning vectors with 3 options for insert excision. The pGEM®-T Vector is ready to use in ligation reactions, prepared by cutting the pGEM®-5Zf(+) Vector with EcoRV and adding a 3´ terminal thymidine to both ends. There was an issue verifying your email address. Our customer and technical support experts are here to help! Trademarks. Contains GoTaq® G2 enzyme. Promega Corporation is a worldwide leader in applying biochemistry and molecular biology to the development of innovative, high-value products for the life sciences. ®Briefly centrifuge the pGEM-T or pGEM®-T Easy Vector and Control Insert … The shoot apex tissues of young seedlings were fixed in RNase-free formalin/acetic acid/alcohol fixative. Promega Corporation is a worldwide leader in applying biochemistry and molecular biology to the development of innovative, high-value products for the life sciences. Literature # TM042. ®Protocol for Ligations Using the pGEM -T and pGEM®-T Easy Vectors and the 2X Rapid Ligation Buffer 3.A. There was an issue creating your account. Molecular Cloning: A Laboratory Manual Third Edition, 1982. This is a free resource for the scientific community that is compiled by Addgene.. Get in touch with a nearby distributor or sales representative. Accordingly, as a means of enhancing tissue invasion, tumor cells use matrix metalloproteinases to degrade ECM proteins. Ligation Using 2X Rapid Ligation Buffer 1. Canada orders: Ship Monday March 15 for arrival on Tuesday March 16. The single major product was cloned into the pGEM-T-easy vector (Promega) and was sequenced. There was an issue creating your account. You have successfully reset your password. Canada orders: Ship Monday March 15 for arrival on Tuesday March 16. One of the easiest methods for cloning blunt-ended DNA fragments including PCR products is T-vector cloning, such as with pGEM®-T or pGEM®-T Easy Vector Systems.This method takes advantage of the “A” overhang added by a PCR enzyme like Taq DNA Polymerase.T vectors are linearized plasmids that have been treated to add 3′ T overhangs to match the A overhangs of the insert. Die In-vitro-Transkription, also die DNA-abhängige Synthese von RNA im Reagenzglas, ist eine molekularbiologische Methode zur Erzeugung von RNA und zur Untersuchung von Promotoren und ihrer Aktivierung durch Transkriptionsfaktoren. The pGEM®-T Easy Vector multiple cloning region is flanked by recognition sites for the restriction enzymes EcoRI, BstZI and NotI, providing three single-enzyme digestions for release of the insert. Shop Now ›, Rapid Ligation for the pGEM®-T and pGEM®-T Easy Vector Systems, Comparing Cloning Efficiency of the pGEM®-T and pGEM®-T Easy Vectors to the TOPO TA Cloning® Vectors, Shorten the Ligation Time for the pGEM®-T Vector Systems, TRE5-A retrotransposition profiling reveals putative RNA polymerase III transcription complex binding sites on the, Privacy Policy and Requests for Information, Insert excision with a BstZI single digest, Ligation can be completed in 1 hour at room temperature, Available with or without competent cells. The Promega mission statement is: To be the most responsive supplier of biological reagents and reagent systems used in research and applied technology applications worldwide. However, the in vivo ECM is comprised not only of proteins but also of a variety of non-protein components. © 2021 Promega Corporation. RNase-resistant, noninfectious virus-like particles containing exogenous RNA sequences (armored RNA) are good candidates as RNA controls and standards in RNA virus detection. (2007) A new 10-min ligation method using a modified buffer system with a very low amount of T4 DNA ligase: the "coffee break ligation" technique. You have not verified your email address. We offer numerous convenient solutions to meet your lab's needs. In the current study, we focused on investigating the mechanisms underlying the development of doxorubicin resistance in osteosarcoma.Methods: The human osteosarcoma cell line MG-63 and doxorubicin-resistant MG-63/Dox cells were used in this study. These polymerases often add a single deoxyadenosine, in a template-independent fashion, to the 3´-ends of the amplified fragments. This paper. The vector allows preparation of single-stranded DNA due to its f1 Origin of Replication. The pGEM®-5Zf(+) Vector serves as a standard cloning vector and as a template for in vitro transcription. US orders: Ship Saturday March 13 for arrival on Monday March 15. These polymerases often add a single deoxyadenosine, in a template-independent fashion, to the 3´-ends of the amplified fragments. Literature # TM042. The Promega mission statement is: To be the most responsive supplier of biological reagents and reagent systems used in research and applied technology applications worldwide. The vectors are prepared by cutting the pGEM®-5Zf(+) and pGEM®-T Easy Vectors, respectively, with EcoR V and adding a 3´ terminal thymidine to both ends. READ PAPER. Privacy Policy and Requests for Information There was an issue logging into your account. Please try again or contact Customer Service. This vector is also known as pGEM®‑5Zf(+). The pGEM-T Vector is prepared by cutting Promega's pGEM-5Zf(+) Vector with EcoR V and adding a 3' terminal thymidine to both ends.These single 3'-T overhangs at the insertion site greatly improve the efficiency of ligation of a PCR product into the plasmid. pGEM-T easy plasmid DNA (500 ng, Promega, Madison, WI, USA) was then added and incubated for 1 h at 37 °C. The position of the T is indicated by * in the pGEM®-T Vector Sequence (.txt). Ratios from 3:1 to 1:3 provide good initial parameters. PDF (548k). To protect your privacy, your account has been locked after 6 failed login attempts. Using the pGEM-T-mH5 vector that we have previously ... Promega) was added and incubated for one additional hour. The vector carries the lacZ alpha-peptide and the multiple cloning region arrangement from pUC18 allowing selection of recombinants by blue/white screening. However, ratios of 8:1 to 1:8 have been used successfully. Download PDF. Yoshino, Y., Ishida, M. and Horii, A. In the pGEM®-T Vector, T7 and SP6 RNA polymerase promoters flank a multiple cloning region within the α-peptide coding region for β-galactosidase. Trademarks. www.promega.com Part# TM042 Printed in USA. The pGEM ®-T and pGEM -T Easy Vector Systems have been optimized using a 1:1 molar ratio of the Control Insert DNA to the vectors. The pGEM®-T Vector was created by linearizing the pGEM®-5Zf(+) Vector with EcoRV at base 51 and adding a T to both 3´-ends. Welcome to Vector Database!. The 12/18 version of this Technical Manual was revised to remove references to discontinued products in the notes on sequencing primers in Section 5.B. There was an issue sending the verification email. The pGEM®-T Vector System II contains JM109 Competent Cells in addition to all of the pGEM®-T Vector System I components. Check your inbox to complete email verification. The pGEM-3Z Vector is intended for use as a standard cloning vector, as well as for the highly efficient synthesis of RNA in vitro. You have successfully reset your password. https://doi.org/10.1530/JME-17-0142 http://jme.endocrinology-journals.org 2018 Society for Endocrinology Printed in Great Britain Published by Bioscientifica Ltd. Please try again or contact Customer Service. The pGEM®-T and pGEM®-T Easy Vector Systems gave a high number of recombinants across a broad range of insert sizes (0.5–3kb) while the TOPO TA Cloning® system worked well for inserts less than 1kb, but showed a striking decrease in performance with larger insert sizes (1–3kb). The pGEM®-11Zf(+) Vector is a standard cloning vector that contains T7 and SP6 RNA polymerase promoters flanking a multiple cloning region within the α-peptide coding region of β-galactosidase. One of the easiest methods for cloning blunt-ended DNA fragments including PCR products is T-vector cloning, such as with pGEM®-T or pGEM®-T Easy Vector Systems.This method takes advantage of the “A” overhang added by a PCR enzyme like Taq DNA Polymerase.T vectors are linearized plasmids that have been treated to add 3′ T overhangs to match the A overhangs of the insert. Dismiss. .. Nicking of DNA was evaluated by ethidium bromide staining after electrophoresis separation in 0.8% agarose gels [ , ]. The pGEM®-3Z Vector is intended for use as a standard cloning vector, as well as for the highly efficient synthesis of RNA in vitro. Frackman, S. and Kephart, D (1999) Rapid ligation for the pGEM®-T and pGEM®-T Easy Vector Systems. The pGEM®-T Easy Vector Systems offer all of the advantages of the pGEM®-T Vector Systems with the added convenience of recognition sites for BstZI, EcoRI and NotI flanking the insertion site. Alternatively, a double digestion may be used to release the insert from the vector. The pGEM®-T Easy Vector Systems offer all of the advantages of the pGEM®-T Vector Systems with the added convenience of recognition sites for BstZI, EcoRI and NotI flanking the insertion site. The pGEM®-T Easy Vector Systems are convenient systems to clone PCR products generated by certain thermostable polymerases. This vector contains T7 and SP6 RNA polymerase promoters flanking a multiple cloning region within the α-peptide coding region of β-galactosidase. Privacy Policy and Requests for Information Note: You will not be able to access your account until your email is verified. PLos ONE, Plate Readers, Fluorometers & Luminometers, Save 20% on pGL4 Luciferase Reporter Vectors, enter PGL20 at checkout. Page 4 Revised 5/07 GGAGA GCTCC CAACG CGTTG GATGC ATAGC TTGAG … Let's find the product that meets your needs. Check your inbox to complete email verification. The pGEM ®-T and pGEM ®-T Easy Vector Systems are convenient systems for the cloning of PCR products.The vectors are prepared by cutting the pGEM ®-5Zf(+) and pGEM ®-T Easy Vectors, respectively, with EcoR V and adding a 3´ terminal thymidine to both ends. The pGEM ®-T and pGEM ®-T Easy Vector Systems are convenient systems for the cloning of PCR products.The vectors are prepared by cutting the pGEM ®-5Zf(+) and pGEM ®-T Easy Vectors, respectively, with EcoR V and adding a 3´ terminal thymidine to both ends. Your password reset link has expired. Description. The pGEM®-T Vector was created by linearizing the pGEM®-5Zf(+) Vector with EcoRV at base 51 and adding a T to both 3´-ends. In addition, we excised the gene encoding GFP-mRNA-96-mer from pTRE-GFP-96-mer and inserted it into plasmid pGEM, because that plasmid contains a bacteriophage T7 promoter. Legal and Trademarks Stay notified of Promega events, products and news. X65308). However, ratios of 8:1 to 1:8 have been used successfully. Our records indicate that this email address is already registered. A short summary of this paper. Download PDF. Please request another reset link. The pGEM®-T Vector Systems are convenient systems to clone PCR products generated by certain thermostable polymerases. Your commerce experience may be limited. Due to our annual Physical inventory orders received after 5PM Central Time on Tuesday March 9 will be shipped according to the following schedule: pGEM®-T Parental vector for TA cloning of PCR products. Revised 12/18 www.promega.com 3. The 3.9-kb product was cloned into pGEM-T Easy (Promega… pGEM®-T Easy Parental vector for TA cloning of PCR products. DNA concentration of linearised recombinant plasmid was determined using the Qubit 1× dsDNA HS Assay Kit (Invitrogen, CA, USA). These polymerases often add a single deoxyadenosine, in a template-independent fashion, to the 3´-ends of the amplified fragments. ... (T. aculeatus and three Zaglossus spp.) As a result, PCR products amplified using GoTaq® DNA Polymerase will contain A-overhangs which makes it suitable for T-vector cloning. The pGEM®-3Zf(+) and pGEM®-3Zf(–) Vectors contain T7 and SP6 RNA polymerase promoters flanking a multiple cloning region within the α-peptide coding region of β-galactosidase. Congratulations! Introduction. The positive samples in this study were termed using the abbreviated name … Promega Notes 71 , 8–9. Legal and Trademarks Download Free PDF. Revised 4/17 www.promega.com 2. Product Components and Storage Conditions PRODUCT SIZE CAT. X65308). Thus, several options exist to remove the desired insert DNA with a single restriction digestion. A verification email has been sent to the primary email address associated with your account. Terms and Conditions Ready-to-use optimized master mix for room-temperature PCR assembly. A password reset email has been sent to the primary email address associated with your account. Please request another reset link. The pGEM®-T and pGEM®-T Easy Vector Systems are convenient systems for the cloning of PCR products. We provide medical information and facilitate research collaborations. Please try again or contact Customer Service. The assay sensitivity was determined using pGEM-T Easy Vector plasmids (Promega, Madison, WI) containing the target sequence of the N (961 bp) and S (1119 bp) genes of SARS-CoV-2. There was an issue resetting your password. Your password reset link has expired. There was an issue resetting your password. Complete Protocol All Rights Reserved. There was an issue verifying your email address. We have successfully cloned PCR products generated using GoTaq® and GoTaq® Flexi DNA Polymerases into the pGEM®-T (Cat.# A3600), pGEM®-T Easy (Cat.# A1360) and pTARGET™ (Cat.# A1410) Vectors. Insertional inactivation of the α-peptide allows recombinant clones to be directly identified by Blue/White Screening on indicator plates. Our records indicate that this email address is already registered. # pGEM®-5Zf(+) Vector 20µg P2241 The pGEM®-5Zf(+) Vector is supplied with a glycerol stock of bacterial strain JM109. Product Components and Storage Conditions PRODUCT SIZE CAT.# pGEM®-7Zf(+) Vector 20µg P2251 The pGEM®-7Zf(+) Vector is provided with a glycerol stock of bacterial strain JM109. If initial experiments with your PCR product are suboptimal, ratio optimization may be necessary. Please contact Customer Service to unlock your account. This page is informational only - this vector is NOT available from Addgene - please contact the manufacturer for further details. Download PDF. 36 Full PDFs related to this paper. There was an issue logging into your account. Thus, several options exist to remove the desired insert DNA with a single restriction digestion. A1360, A1380, A3600, A3610. Enter your username and we'll send a link to reset your password. Download Full PDF Package. A3600. By creating an account, you confirm that you accept the, Plate Readers, Fluorometers & Luminometers, Privacy Policy and Requests for Information. Our customer and technical support experts are here to help! The pGEM®-T Vector is ready to use in ligation reactions, prepared by cutting the pGEM®-5Zf(+) Vector with EcoRV and adding a 3´ terminal thymidine to both ends. In this study, we describe a method for producing armored L-RNA. The gold arrows indicate 5′-RACE products characterized from the Δ5G construct. pGEM®-T Easy Vector Systemは、従来のpGEM®-T Vector Systemの機能に加え、マルチクローニングサイトの両端にEcoRIとNotIサイトが加えられました。そのため、1種類（NotI、EcoRIあるいはBstZI）の制限酵素を用いるだけで、クローニング後のインサートDNAを簡単に切り出すことがきます。 Please try again or contact Customer Service. Instructions for Use of Product(s) Reactions using this buffer may be incubated for 1 hour at room temperature. The insertion site is flanked by BstZI sites. We've detected that you are using an older version of Internet Explorer. In this study, a specific 277-bp cDNA fragment of DHD4 was amplified and then cloned into the pGEM-T Easy vector (Promega), which was used to produce antisense and sense RNA probes. The multiple cloning site is flanked by recognition sites for the restriction enzyme BstZI, allowing release of the insert by a single-enzyme digestion. Summary of Changes Get in touch with a nearby distributor or sales representative. To protect your privacy, your account will be locked after 6 failed attempts. Our website uses functional cookies that do not collect any personal information or track your browsing activity. The pGEM®-T Vector is derived from the pGEM®-5Zf(+) Vector (GenBank® Accession No. Second-generation, high-performance GoTaq® G2 DNA Polymerase with Mg-free buffers. A verified email address is required to access the full functionality of your Promega.com account. By creating an account, you confirm that you accept the, pGEM®-T and pGEM®-T Easy Vector Systems Technical Manual, pGEM T and pGEM T Easy Vector Systems FB033, 2017
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