All Rights Reserved. However, ratios of 8:1 to 1:8 have been used successfully. READ PAPER. You have not verified your email address. pGEM®-T Easy Parental vector for TA cloning of PCR products. Quick PROTOCOL 1 pGEM®-T and pGEM®-T Easy Vector Systems INSTRUCTIONS FOR USE OF PRODUCTS A1360, A1380, A3600 AND A3610. There was an issue logging into your account. Please try again or contact Customer Service. Main. Revised 4/17 www.promega.com 2. Our website uses functional cookies that do not collect any personal information or track your browsing activity. Terms and Conditions Accordingly, as a means of enhancing tissue invasion, tumor cells use matrix metalloproteinases to degrade ECM proteins. Our website uses functional cookies that do not collect any personal information or track your browsing activity. A verification email has been sent to the primary email address associated with your account. Download PDF. Congratulations! The incubation period may be extended to increase the number of colonies after transformation. Due to our annual Physical inventory orders received after 5PM Central Time on Tuesday March 9 will be shipped according to the following schedule: All Rights Reserved. Get in touch with a nearby distributor or sales representative. 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. 36 Full PDFs related to this paper. Download Free PDF. Literature # TM042. The pGEM®-T Vector is derived from the pGEM®-5Zf(+) Vector (GenBank® Accession No. A verified email address is required to access the full functionality of your Promega.com account. 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. 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 The extracellular matrix (ECM) plays an important role in maintaining tissue homeostasis and poses a significant physical barrier to in vivo cell migration. Note: You will not be able to access your account until your email is verified. Promega Corporation is a worldwide leader in applying biochemistry and molecular biology to the development of innovative, high-value products for the life sciences. Page 4 Revised 5/07 GGAGA GCTCC CAACG CGTTG GATGC ATAGC TTGAG … The position of the T is indicated by * in the pGEM®-T Vector Sequence (.txt). 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. PDF (548k). Please try again or contact Customer Service. Please update your browser to Internet Explorer 11 or above. In this study, we describe a method for producing armored L-RNA. PCR products with low concentration or generating heterogeneity in the sequencing chromatograms were cloned into pGEM-T Easy Vector (Promega) for sequencing. www.promega.com Part# TM042 Printed in USA. Due to our annual Physical inventory orders received after 5PM Central Time on Tuesday March 9 will be shipped according to the following schedule: There was an issue verifying your email address. Please try again or contact Customer Service. We provide medical information and facilitate research collaborations. 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. There was an error processing your request. Ratios from 3:1 to 1:3 provide good initial parameters. 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. 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. Reactions using this buffer may be incubated for 1 hour at room temperature. Thank you for verifying your email address. X65308). Please check your network settings and try again. PCR cloning system for expression in mammalian cells. 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. Thank you for verifying your email address. Catalog number selected: US orders: Ship Saturday March 13 for arrival on Monday March 15. See Protocol for detailed storage recommendations. 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. 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. Product Components and Storage Conditions PRODUCT SIZE CAT. Please try again or contact Customer Service. Insertional inactivation of the α-peptide allows recombinant clones to be directly identified by Blue/White Screening on indicator plates. Description. Enter your username and we'll send a link to reset your password. Vector database is a digital collection of vector backbones assembled from publications and commercially available sources. Frackman, S. and Kephart, D (1999) Rapid ligation for the pGEM®-T and pGEM®-T Easy Vector Systems. The pGEM®-T Vector is derived from the pGEM®-5Zf(+) Vector (GenBank® Accession No. 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. Check your inbox to complete email verification. A password reset email has been sent to the primary email address associated with your account. You have not verified your email address. Let's find the product that meets your needs. Our customer and technical support experts are here to help! 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. Alternatively, a double digestion may be used to release the insert from the vector. Legal and Trademarks The vector carries the lacZ alpha-peptide and the multiple cloning region arrangement from pUC18 allowing selection of recombinants by blue/white screening. The pGEM®-T Easy Vector Systems are convenient systems to clone PCR products generated by certain thermostable polymerases. The single major product was cloned into the pGEM-T-easy vector (Promega) and was sequenced. PLos ONE, Plate Readers, Fluorometers & Luminometers, Save 20% on pGL4 Luciferase Reporter Vectors, enter PGL20 at checkout. After that, you will need to contact Customer Service to unlock your account. There was an issue creating your account. (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. Ligation Protocol 1. PCR products were gel-purified, cloned into the pGEM-T Easy Vector system (Promega Corporation, WI, USA) and sequenced using the BigDye Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems). You've created a Promega.com account. The vector allows preparation of single-stranded DNA due to its f1 Origin of Replication. The pGEM®-T Vector Systems are convenient systems to clone PCR products generated by certain thermostable polymerases. © 2021 Promega Corporation. There was an issue with the password reset process. Check your inbox to complete email verification. Please request another reset link. To protect your privacy, your account will be locked after 6 failed attempts. As a result, PCR products amplified using GoTaq® DNA Polymerase will contain A-overhangs which makes it suitable for T-vector cloning. There was an issue sending the verification email. Instructions for Use of Product(s) A1360, A1380, A3600, A3610. Stay notified of Promega events, products and news. A short summary of this paper. There was an issue sending the verification email. ... (T. aculeatus and three Zaglossus spp.) 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. The insertion site is flanked by BstZI, EcoRI, and NotI sites. To protect your privacy, your account has been locked after 6 failed login attempts. Insertional inactivation of the alpha-peptide allows recombinant clones to be directly identified by blue/white screening on indicator plates. The pGEM®-T Vector cloning region is flanked by recognition sites for the enzyme BstZI. Our records indicate that this email address is already registered. Plates were developed using 1 … The position of the T is indicated by * in the pGEM®-T Vector Sequence (.txt). Download Full PDF Package. This is a free resource for the scientific community that is compiled by Addgene.. DNA concentration of linearised recombinant plasmid was determined using the Qubit 1× dsDNA HS Assay Kit (Invitrogen, CA, USA). pGEM®-T Parental vector for TA cloning of PCR products. 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. Please try again or contact Customer Service. Please try again or contact Customer Service. Your password reset link has expired. However, ratios of 8:1 to 1:8 have been used successfully. Our customer and technical support experts are here to help! Issai Falcon. The pGEM®-T and pGEM®-T Easy Vector Systems are convenient systems for the cloning of PCR products. Trademarks. 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. You've created a Promega.com account. The insertion site is flanked by BstZI sites. When you select your country, you agree that we can place these functional cookies on your device. Molecular Cloning: A Laboratory Manual Third Edition. Please try again or contact Customer Service. The multiple cloning site is flanked by recognition sites for the restriction enzyme BstZI, allowing release of the insert by a single-enzyme digestion. However, the in vivo ECM is comprised not only of proteins but also of a variety of non-protein components. 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. Briefly centrifuge the pGEM ®-T or pGEM -T Easy Vector and Control Insert DNA tubes to collect contents at the bottom of the tube. 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. These single 3´-T overhangs at the insertion site greatly improve the efficiency of ligation of a PCR product into the plasmid by preventing recircularization of the vector and providing a compatible overhang for ligation of PCR products with A overhangs. Revised 12/18 www.promega.com 3. To protect your privacy, your account has been locked after 6 failed login attempts. This vector is also known as pGEM®‑5Zf(+). Instructions for Use of Product(s) To protect your privacy, your account will be locked after 6 failed attempts. Please try again or contact Customer Service. The pGEM®-T and pGEM®-T Easy Vector Systems include a 2X Rapid Ligation Buffer for ligation of PCR products. Second-generation, high-performance GoTaq® G2 DNA Polymerase with Mg-free buffers. These samples were collected from Da Longshu village (a, 45 samples), Bai Shiyan village (b, 28 … 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. Using the pGEM-T-mH5 vector that we have previously ... Promega) was added and incubated for one additional hour. The positive samples in this study were termed using the abbreviated name … Thus, several options exist to remove the desired insert DNA with a single restriction digestion. Contains GoTaq® G2 enzyme. Thus, several options exist to remove the desired insert DNA with a single restriction digestion. Ligation Using 2X Rapid Ligation Buffer 1. This paper. 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. Instructions for Use of Product(s) A1360, A1380, A3600, A3610. 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. Your password reset link has expired. There was an issue logging into your account. Trademarks. Please try again or contact Customer Service. We evaluated the cloning efficiency of different size PCR products into three T-vector cloning systems. Please request another reset link. Revised 4/17 www.promega.com 2. 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. There was an issue resetting your password. This product is available through the Promega Helix onsite stocking program. Please try again or contact Customer Service. However, the length of RNA packaged in the virus-like particles with high efficiency is usually less than 500 bases. .. Nicking of DNA was evaluated by ethidium bromide staining after electrophoresis separation in 0.8% agarose gels [ , ]. Our records indicate that this email address is already registered. Dismiss. Privacy Policy and Requests for Information The pGEM®-5Zf(+) Vector serves as a standard cloning vector and as a template for in vitro transcription. If initial experiments with your PCR product are suboptimal, ratio optimization may be necessary. We provide medical information and facilitate research collaborations. There was an issue with the password reset process. Canada orders: Ship Monday March 15 for arrival on Tuesday March 16. PCR cloning vectors with 3 options for insert excision. Introduction. After that, you will need to contact Customer Service to unlock your account. RNase-resistant, noninfectious virus-like particles containing exogenous RNA sequences (armored RNA) are good candidates as RNA controls and standards in RNA virus detection. © 2021 Promega Corporation. A password reset email has been sent to the primary email address associated with your account. A resource designed for scientists just embarking on their career, focusing on fundamental technologies and techniques. A verified email address is required to access the full functionality of your Promega.com account. 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. Congratulations! You have successfully reset your password. Please check your network settings and try again. Your commerce experience may be limited. Download PDF. Diese Seite wurde zuletzt am … A verification email has been sent to the primary email address associated with your account. Get in touch with a nearby distributor or sales representative. The shoot apex tissues of young seedlings were fixed in RNase-free formalin/acetic acid/alcohol fixative. Please try again or contact Customer Service. Promega Notes 71 , 8–9. 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 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. 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. These polymerases often add a single deoxyadenosine, in a template-independent fashion, to the 3´-ends of the amplified fragments. Background: Development of resistance to doxorubicin-based chemotherapy limits its curative effect in osteosarcoma. This page is informational only - this vector is NOT available from Addgene - please contact the manufacturer for further details. You have successfully reset your password. 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. Literature # TM042. Molecular Cloning: A Laboratory Manual Third Edition, 1982. We prepared a series of pGEM plasmids (Promega) containing 1, 2, 4, 8, 16, 32, or 64 tandem repeats of the sequence described above. The pGEM®-T Vector System II contains JM109 Competent Cells in addition to all of the pGEM®-T Vector System I components. If initial experiments with your PCR product are suboptimal, ratio optimization may be necessary. 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. A1360, A1380, A3600, A3610. There was an issue creating your account. These polymerases often add a single deoxyadenosine, in a template-independent fashion, to the 3´-ends of the amplified fragments. Abstract. Please contact Customer Service to unlock your account. 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. Stay notified of Promega events, products and news. Terms and Conditions Complete Protocol We offer numerous convenient solutions to meet your lab's needs. 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.

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