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Pcr buffer 1

Manufactured by Roche

PCR buffer 1 is a liquid solution designed to facilitate the polymerase chain reaction (PCR) process. It provides the necessary components, such as ions and cofactors, to support the optimal performance of the DNA polymerase enzyme during the amplification of DNA sequences.

Automatically generated - may contain errors

2 protocols using pcr buffer 1

1

Single-Cell Polylox Sequencing Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols
For the isolation of single HSC, BM cells were stained as described
above. During FACS sorting, individual cells were deposited into 8-tube PCR
stripes containing in each well 25 μl lysis buffer (12.6 μg
proteinase K (Thermo Fisher Scientific) in PCR buffer 1 (Roche)). Lysis was done
for 1 h at 55 °C, terminated at 95 °C for 10 min, and cooled to 4
°C before adding the remaining PCR reagents to a final volume of 50
μl. The Polylox cassette was then amplified by nested
PCR. First round PCR: primer #2450 (see above) and primer #494
(5′-AGCTACAGCCTCGATTTGTGGTG-3′) for 5 min at 95 °C; (30 s
at 95 °C, 30 s at 56 °C, 5 min at 72 °C) 35 times; 10 min
at 72 °C. Second round PCR: 1-2 μl of first PCR reaction was used
as template and amplified with primers #2426
5′-CGACGACACTGCCAAAGATTTC-3′ and #2427 (see above) for 5 min at 95
°C; (30 s at 95 °C, 30 s at 62 °C, 5 min at 72 °C)
35 times; 10 min at 72 °C. The nested PCR products were purified by
QIAquick PCR Purification Kit (Qiagen), analyzed by gel electrophoresis for
product length, and analyzed by Sanger sequencing (GATC Biotech). Barcodes were
decoded for each of these sequences.
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2

Single-Cell Polylox Sequencing Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols
For the isolation of single HSC, BM cells were stained as described
above. During FACS sorting, individual cells were deposited into 8-tube PCR
stripes containing in each well 25 μl lysis buffer (12.6 μg
proteinase K (Thermo Fisher Scientific) in PCR buffer 1 (Roche)). Lysis was done
for 1 h at 55 °C, terminated at 95 °C for 10 min, and cooled to 4
°C before adding the remaining PCR reagents to a final volume of 50
μl. The Polylox cassette was then amplified by nested
PCR. First round PCR: primer #2450 (see above) and primer #494
(5′-AGCTACAGCCTCGATTTGTGGTG-3′) for 5 min at 95 °C; (30 s
at 95 °C, 30 s at 56 °C, 5 min at 72 °C) 35 times; 10 min
at 72 °C. Second round PCR: 1-2 μl of first PCR reaction was used
as template and amplified with primers #2426
5′-CGACGACACTGCCAAAGATTTC-3′ and #2427 (see above) for 5 min at 95
°C; (30 s at 95 °C, 30 s at 62 °C, 5 min at 72 °C)
35 times; 10 min at 72 °C. The nested PCR products were purified by
QIAquick PCR Purification Kit (Qiagen), analyzed by gel electrophoresis for
product length, and analyzed by Sanger sequencing (GATC Biotech). Barcodes were
decoded for each of these sequences.
+ Open protocol
+ Expand

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