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Purelink pcr micro kit

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The PURELINK™ PCR Micro Kit is a DNA purification kit designed for the rapid and efficient extraction of PCR amplicons from small-scale reactions. It utilizes a simple spin-column format to isolate and purify PCR products from reaction mixtures.

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Lab products found in correlation

Kapa hifi hotstart readymix, by Roche (55 mentions) Agilent 2100 bioanalyzer, by Agilent Technologies (3 mentions) Solid total rna seq kit, by Thermo Fisher Scientific (3 mentions) Solid 4 slide, by Thermo Fisher Scientific (2 mentions) Qubit fluorometer, by Thermo Fisher Scientific (2 mentions) Micropoly a purist kit, by Thermo Fisher Scientific (1 mentions) Megaclear kit, by Thermo Fisher Scientific (1 mentions) T7 rna polymerase, by New England Biolabs (1 mentions) Phusion polymerase, by New England Biolabs (1 mentions) Agilent 2100, by Agilent Technologies (1 mentions) Rnaclean xp beads, by Beckman Coulter (1 mentions) Miseq 300, by Illumina (1 mentions) Qubit spectrometer, by Thermo Fisher Scientific (1 mentions) Bioanalyzer, by Agilent Technologies (1 mentions) Q5 polymerase, by New England Biolabs (1 mentions) Lysostaphin, by Merck Group (1 mentions) Instagene matrix, by Bio-Rad (1 mentions) Directpcr cell lysis buffer, by Viagen Biotech (1 mentions) X tremegene 9, by Merck Group (1 mentions) T7 endonuclease 1 t7e1, by New England Biolabs (1 mentions)

Spelling variants of this product

PureLink kit (67 citations) Purelink Micro Kit (7 citations)

72 protocols using purelink pcr micro kit

1

Reverse Transcription and cDNA Preparation

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Example 2

PCR procedures for the preparation of cDNA are performed using 2×KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, Mass.). This system includes 2×KAPA ReadyMix12.5 μl; Forward Primer (10 uM) 0.75 μl; Reverse Primer (10 uM) 0.75 μl; Template cDNA 100 ng; and dH2O diluted to 25.0 The reaction conditions are at 95° C. for 5 min. and 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min. then 4° C. to termination.

The reverse primer of the instant invention incorporates a poly-T120 for a poly-A120 in the mRNA. Other reverse primers with longer or shorter poly(T) tracts can be used to adjust the length of the poly(A) tail in the mRNA.

The reaction is cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, Calif.) per manufacturer's instructions (up to 5 μg). Larger reactions will require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA is quantified using the NANODROP™ and analyzed by agarose gel electrophoresis to confirm the cDNA is the expected size. The cDNA is then submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US10493167B2. In vivo production of proteins (2019-12-03). ModernaTX, Inc. [US]. Inventors: Antonin de Fougerolles [BE], Justin Guild [US].
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2

cDNA Preparation and Cleanup Protocol

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Example 2

PCR procedures for the preparation of cDNA can be performed using 2×KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, MA). This system includes 2×KAPA ReadyMix12.5 μl; Forward Primer (10 μM) 0.75 μl; Reverse Primer (10 μM) 0.75 μl; Template cDNA—100 ng; and dH2O diluted to 25.0 μl. The PCR reaction conditions can be: at 95° C. for 5 min. and 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min. then 4° C. to termination.

The reverse primer of the instant invention can incorporate a poly-T120 for a poly-A120 in the mRNA. Other reverse primers with longer or shorter poly(T) tracts can be used to adjust the length of the poly(A) tail in the polynucleotide mRNA.

The reaction can be cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, CA) per manufacturer's instructions (up to 5 μg). Larger reactions will require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA can be quantified using the NANODROP™ and analyzed by agarose gel electrophoresis to confirm the cDNA is the expected size. The cDNA can then be submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US11802146B2. Polynucleotides encoding anti-chikungunya virus antibodies (2023-10-31). ModernaTX, Inc. [US], Vanderbilt University [US]. Inventors: Sunny Himansu [US], James E. Crowe, Jr. [US], Giuseppe Ciaramella [US].
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3

cDNA Preparation and Characterization

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Example 2

PCR procedures for the preparation of cDNA can be performed using 2×KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, Mass.). This system includes 2×KAPA ReadyMix12.5 μl; Forward Primer (10 μM) 0.75 μl; Reverse Primer (10 μM) 0.75 μl; Template cDNA—100 ng; and dH2O diluted to 25.0 μl. The PCR reaction conditions can be: at 95° C. for 5 min. and 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min. then 4° C. to termination.

The reverse primer of the instant disclosure can incorporate a poly-T120 for a poly-A120 in the mRNA. Other reverse primers with longer or shorter poly(T) tracts can be used to adjust the length of the poly(A) tail in the polynucleotide mRNA.

The reaction can be cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, Calif.) per manufacturer's instructions (up to 5 μg). Larger reactions will require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA can be quantified using the NANODROP™ and analyzed by agarose gel electrophoresis to confirm the cDNA is the expected size. The cDNA can then be submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US11504337B2. Polynucleotides encoding methylmalonyl-CoA mutase (2022-11-22). ModernaTX, Inc. [US]. Inventors: Paolo Martini [US], Vladimir Presnyak [US], Kerry Benenato [US], Stephen Hoge [US], Iain McFadyen [US], Ellalahewage Sathyajith Kumarasinghe [US].
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4

cDNA Preparation and Sequencing Protocol

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Example 3

PCR procedures for the preparation of cDNA are performed using 2×KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, Mass.). This system includes 2×KAPA ReadyMix12.5 μl; Forward Primer (10 uM) 0.75 μl; Reverse Primer (10 uM) 0.75 μl; Template cDNA—100 ng; and dH2O diluted to 25.0 μl. The reaction conditions are at 95° C. for 5 min. and 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min. then 4° C.

The reaction is cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, Calif.) per manufacturer's instructions (up to 5 μg). Larger reactions will require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA is quantified using the NANODROP™ and analyzed by agarose gel electrophoresis to confirm the cDNA is the expected size. The cDNA is then submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US11202793B2. High purity RNA compositions and methods for preparation thereof (2021-12-21). ModernaTX, Inc. [US]. Inventors: Stephen Hoge [US], William Issa [US], Edward J. Miracco [US], Jennifer Nelson [US], Amy E. Rabideau [US], Gabor Butora [US].
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5

cDNA Preparation and Purification Protocol

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Example 3

PCR procedures for the preparation of cDNA are performed using 2×KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, Mass.). This system includes 2×KAPA ReadyMix 12.5 μl; Forward Primer (10 μM) 0.75 μl; Reverse Primer (10 μM) 0.75 μl; Template cDNA —100 ng; and dH20 diluted to 25.0 μl. The reaction conditions are at 95° C. for 5 min. and 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min. then 4° C. to termination.

The reaction is cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, Calif.) per manufacturer's instructions (up to 5 μg). Larger reactions will require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA is quantified using the NANODROP™ and analyzed by agarose gel electrophoresis to confirm the cDNA is the expected size. The cDNA is then submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US11541113B2. Human cytomegalovirus vaccine (2023-01-03). ModernaTX, Inc. [US]. Inventors: Giuseppe Ciaramella [US], Shinu John [US].
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6

Whole Transcriptome Profiling of S1P and VM Cells

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Whole transcriptome libraries from two S1P pools and two VM replicates were produced using MicroPolyA Purist Kit (Ambion) and SOLiD Total RNA-Seq kit (Applied Biosystems). Briefly, approximately 200 ng/sample poly (A)-positive RNA was enriched using MicroPoly (A) Purist Kit from 15–20 μg total RNA. Quality and concentration of the extracted poly (A)-positive RNA was re-assessed as described above, and poly (A)-positive RNA was digested with RNase III. Ligation of the adaptor mix and reverse transcription was performed following the manufacturer instructions. cDNA libraries were size selected for 150–250 bp fragments, amplified in 15–18 PCR cycles using barcoded adaptor primers and purified with PureLink PCR micro kit (Invitrogen). These barcoded cDNA libraries were then amplified by emulsion PCR from 0.5 pM template. Sequencing beads from the barcoded libraries were pooled and loaded on a SOLiD 4 slide (Applied Biosystems), according to manufacturer’s instructions. SOLiD ToP Sequencing chemistry was used to produce pair end (50 bp + 35 bp) sequencing reads.
Plaza D.F., Lin C.W., van der Velden N.S., Aebi M, & Künzler M. (2014). Comparative transcriptomics of the model mushroom Coprinopsis cinerea reveals tissue-specific armories and a conserved circuitry for sexual development. BMC Genomics, 15(1), 492.
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7

Small RNA-Seq Library Preparation Protocol

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20 ng of EV RNA and 120 ng cell RNA, both enriched for small RNA, was used as the starting input for RNA-Seq library preparation. The preparation of the cDNA library was done according to the protocol from Applied Biosystems SOLiD™ small RNA library preparation from Invitrogen. The cDNA fragments were barcoded in the PCR amplification step to enable simultaneous sequencing of different samples in a single run. The cDNAs underwent 18 cycles of PCR using barcoded primers. The PCR products were purified using PureLink™ PCR Micro Kit (Invitrogen) and analyzed for size and concentration on Agilent 2100 Bioanalyzer using DNA HS chips. Equal molar amount of each barcoded sample were pooled together in one library, which subsequently were used in emulsion PCR to a total concentration of 0.5 pM. Approximately 1 billion enriched beads were deposited on a full glass slide for SOLiD 5500xl sequencing. Cell RNA from Hs578T and AU565 was previously sequenced on SOLiD4 [42 ], SRA accession SRX273660 (AU565) and SRX273750 (Hs578T). All sequencing was performed at the genomic facility, Nord University. Raw sequences were submitted to the National Center for Biotechnology Information (NCBI) Short Read Archive, Bioproject PRJNA309295
Fiskaa T., Knutsen E., Nikolaisen M.A., Jørgensen T.E., Johansen S.D., Perander M, & Seternes O.M. (2016). Distinct Small RNA Signatures in Extracellular Vesicles Derived from Breast Cancer Cell Lines. PLoS ONE, 11(8), e0161824.
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8

cDNA Synthesis and Purification

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Example 2

PCR procedures for the preparation of cDNA are performed using 2× KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, Mass.). This system includes 2× KAPA ReadyMix 12.5 μl; Forward Primer (10 uM) 0.75 μl; Reverse Primer (10 uM) 0.75 μl; Template cDNA 100 ng; and dH20 diluted to 25.0 μl. The reaction conditions are at 95° C. for 5 min. and 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min. then 4° C. to termination.

The reverse primer of the instant invention incorporates a poly-T120 for a poly-A120 in the mRNA. Other reverse primers with longer or shorter poly(T) tracts can be used to adjust the length of the poly(A) tail in the mRNA.

The reaction is cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, Calif.) per manufacturer's instructions (up to 5 μg). Larger reactions will require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA is quantified using the NANODROP™ and analyzed by agarose gel electrophoresis to confirm the cDNA is the expected size. The cDNA is then submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US10077439B2. Removal of DNA fragments in mRNA production process (2018-09-18). ModernaTX, Inc. [US]. Inventors: William Joseph Issa [US], Yuxun Wang [US], Stephane Bancel [US].
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9

cDNA Synthesis and Purification Protocol

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Example 3

PCR procedures for the preparation of cDNA may be performed using 2×KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, Mass.). This system includes 2×KAPA ReadyMix 12.5 μl; Forward Primer (10 μM) 0.75 μl; Reverse Primer (10 μM) 0.75 μl; Template cDNA 100 ng; and dH2O diluted to 25.0 μl. The reaction conditions may be at 95° C. for 5 min. The reaction may be performed for 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min, then 4° C. to termination.

The reaction may be cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, Calif.) per manufacturer's instructions (up to 5 μg). Larger reactions may require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA may be quantified using the NANODROP™ and analyzed by agarose gel electrophoresis to confirm that the cDNA is the expected size. The cDNA may then be submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US11278611B2. Zika virus RNA vaccines (2022-03-22). ModernaTX, Inc. [US]. Inventors: Giuseppe Ciaramella [US], Sunny Himansu [US], Eric Yi-Chun Huang [US], Tal Zaks [US].
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10

Poly(A) Tail cDNA Synthesis Protocol

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Example 2

PCR procedures for the preparation of cDNA are performed using 2×KAPA HIFI™ HotStart ReadyMix by Kapa Biosystems (Woburn, Mass.). This system includes 2×KAPA ReadyMix12.5 μl; Forward Primer (10 uM) 0.75 μl; Reverse Primer (10 uM) 0.75 μl; Template cDNA 100 ng; and dH2O diluted to 25.0 μl. The reaction conditions are at 95° C. for 5 min. and 25 cycles of 98° C. for 20 sec, then 58° C. for 15 sec, then 72° C. for 45 sec, then 72° C. for 5 min. then 4° C. to termination.

The reverse primer of the instant invention incorporates a poly-T120 for a poly-A120 in the mRNA. Other reverse primers with longer or shorter poly(T) tracts can be used to adjust the length of the poly(A) tail in the mRNA.

The reaction is cleaned up using Invitrogen's PURELINK™ PCR Micro Kit (Carlsbad, Calif.) per manufacturer's instructions (up to 5 μg). Larger reactions will require a cleanup using a product with a larger capacity. Following the cleanup, the cDNA is quantified using the NanoDrop and analyzed by agarose gel electrophoresis to confirm the cDNA is the expected size. The cDNA is then submitted for sequencing analysis before proceeding to the in vitro transcription reaction.

US10898574B2. Delivery and formulation of engineered nucleic acids (2021-01-26). ModernaTX, Inc. [US]. Inventors: Antonin de Fougerolles [BE], Sayda M. Elbashir [US].
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