BioMicroCenter:PippinPrep - Revision history

Hilo1: /* Preparative Gel Electrophoresis */

2025-04-25T17:20:31Z

Preparative Gel Electrophoresis

← Older revision		Revision as of 17:20, 25 April 2025
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	\|}		\|}
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	The BluePippin (Sage Science) is an automated preparative gel electrophoresis system that simplifies size fractionation of DNA. Runs of the five lane gels take approximately 1 hour to overnight depending on the fragment length and can either use internal/internal lane markers for calibration or ethidium bromide to detect passing bands. The BluePippin can separate bands between 50bp and 50kb and utilizes pulse field electrophoresis to separate larger fragment sizes. The Pippin Prep is meant for shorter fragments with a range from 100bp – 1.5kb~~. We have done some testing of the BluePippin system and the results are [[BioMicroCenter:PippinPrep_Testing\|'''HERE''']]~~. <BR><BR>		The BluePippin (Sage Science) is an automated preparative gel electrophoresis system that simplifies size fractionation of DNA. Runs of the five lane gels take approximately 1 hour to overnight depending on the fragment length and can either use internal/internal lane markers for calibration or ethidium bromide to detect passing bands. The BluePippin can separate bands between 50bp and 50kb and utilizes pulse field electrophoresis to separate larger fragment sizes. The Pippin Prep is meant for shorter fragments with a range from 100bp – 1.5kb. <BR><BR>

	Critically, both systems require large amounts of input for larger fragment sizes. Yields of <50% are typical '''within the desired range''' so multiple ug of input are strongly recommended before using either Pippin system. Samples with lower amounts of input should consider alternative approaches including SPRIbead size selection.<BR><BR>		Critically, both systems require large amounts of input for larger fragment sizes. Yields of <50% are typical '''within the desired range''' so multiple ug of input are strongly recommended before using either Pippin system. Samples with lower amounts of input should consider alternative approaches including SPRIbead size selection.<BR><BR>

Hilo1 at 17:19, 25 April 2025

2025-04-25T17:19:27Z

← Older revision		Revision as of 17:19, 25 April 2025
Line 2:		Line 2:


	The ~~BioMicroCenter~~ utilizes the Sage Scientific Blue Pippin and the Pippin Prep for any gel based sizing. The instrument can be ~~run as a walk-up~~ or as an assisted service as part of ~~Illumina, PacBio, or Oxford Nanopore~~ library prep. <BR><BR><BR>		The BioMicro Center utilizes the Sage Scientific Blue Pippin and the Pippin Prep for any gel based sizing. The instrument can be scheduled through iLabs or as an assisted service as part of any NGS library prep in the Center. <BR><BR><BR>

	== Preparative Gel Electrophoresis ==		== Preparative Gel Electrophoresis ==

Hilo1: /* Preparative Gel Electrophoresis */

2025-04-25T17:18:42Z

Preparative Gel Electrophoresis

← Older revision		Revision as of 17:18, 25 April 2025
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	\|2.0%		\|2.0%
	\|100-600bp		\|100-600bp
	\|		\|Specific size libraries
	\|-		\|-
	\|1.5%		\|1.5%
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	\|-		\|-
	\|0.75%		\|0.75%
	\|~~1.0~~-50kb		\|1kb-50kb
	\|~~Uses pulse field.~~		\|Long read
	\|}		\|}
	\|}		\|}

Hilo1: /* Preparative Gel Electrophoresis */

2025-04-25T17:17:04Z

Preparative Gel Electrophoresis

← Older revision		Revision as of 17:17, 25 April 2025
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	\|		\|
	[[Image:BMC_Pippin.jpg\|thumb\|right\|250px\|BluePippin]]		[[Image:BMC_Pippin.jpg\|thumb\|right\|250px\|BluePippin]]
	[[Image:BMC_pippinGel.~~png~~\|thumb\|right\|250px\|~~BluePippin Gel~~]]		[[Image:BMC_pippinGel.jpg\|thumb\|right\|250px\|PippinPrep]]

	\|- style="vertical-align: top;"		\|- style="vertical-align: top;"

Hilo1 at 17:15, 25 April 2025

2025-04-25T17:15:47Z

← Older revision		Revision as of 17:15, 25 April 2025
Line 2:		Line 2:


	The BioMicroCenter utilizes the Sage Scientific Blue Pippin ~~;http://www.sagescience.com/wp-content/uploads/2016/09/BluePippin-Operations-Manual-460013-Rev-D.pdf]~~ for any gel based sizing. The instrument can be run as a walk-up or as an assisted service as part of Illumina, PacBio, or Oxford Nanopore library prep. <BR><BR><BR>		The BioMicroCenter utilizes the Sage Scientific Blue Pippin and the Pippin Prep for any gel based sizing. The instrument can be run as a walk-up or as an assisted service as part of Illumina, PacBio, or Oxford Nanopore library prep. <BR><BR><BR>

	== Preparative Gel Electrophoresis ==		== Preparative Gel Electrophoresis ==
Line 28:		Line 28:
	\|}		\|}
	\|		\|
	The BluePippin (Sage Science) is an automated preparative gel electrophoresis system that simplifies size fractionation of DNA. Runs of the five lane gels take approximately 1 hour to overnight depending on the fragment length and can either use internal/internal lane markers for calibration or ethidium bromide to detect passing bands. The BluePippin can separate bands between 50bp and 50kb. We have done some testing of the BluePippin system and the results are [[BioMicroCenter:PippinPrep_Testing\|'''HERE''']]. <BR><BR>		The BluePippin (Sage Science) is an automated preparative gel electrophoresis system that simplifies size fractionation of DNA. Runs of the five lane gels take approximately 1 hour to overnight depending on the fragment length and can either use internal/internal lane markers for calibration or ethidium bromide to detect passing bands. The BluePippin can separate bands between 50bp and 50kb and utilizes pulse field electrophoresis to separate larger fragment sizes. The Pippin Prep is meant for shorter fragments with a range from 100bp – 1.5kb. We have done some testing of the BluePippin system and the results are [[BioMicroCenter:PippinPrep_Testing\|'''HERE''']]. <BR><BR>

	Critically, ~~the system does~~ require large amounts of input. Yields of <50% are typical '''within the desired range''' so multiple ug of input are strongly recommended before using ~~the~~ Pippin ~~systems~~. Samples with lower amounts of input should consider alternative approaches including SPRIbead ~~cuts~~.<BR><BR>		Critically, both systems require large amounts of input for larger fragment sizes. Yields of <50% are typical '''within the desired range''' so multiple ug of input are strongly recommended before using either Pippin system. Samples with lower amounts of input should consider alternative approaches including SPRIbead size selection.<BR><BR>

	The system is based on precast gel cartridges. These gels narrow as they run, concentrating the sample from the initial 40ul (30ul sample + 10ul loading buffer) loaded in the wells. The BluePippin uses an optical detector to measure the rate of passage of bands based on high speed fluorescent markers or by directly detecting the DNA using ethidium bromide. Once the desired band passes the detector, the valve on each lane switches from a waste reservoir to an elution chamber. Once the band is passed, the valve turns back to waste and the desired band can be removed from the elution chamber with a pippette. Samples can be cleaned up with either SPRI beads or using an ethanol precipitation.		The system is based on precast gel cartridges. These gels narrow as they run, concentrating the sample from the initial 40ul (30ul sample + 10ul loading buffer) loaded in the wells. The BluePippin uses an optical detector to measure the rate of passage of bands based on high speed fluorescent markers or by directly detecting the DNA using ethidium bromide. Once the desired band passes the detector, the valve on each lane switches from a waste reservoir to an elution chamber. Once the band is passed, the valve turns back to waste and the desired band can be removed from the elution chamber with a pippette. Samples can be cleaned up with either SPRI beads or using an ethanol precipitation.
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	\|- style="vertical-align: top;"		\|- style="vertical-align: top;"
	\| colspan="2" \|		\| colspan="2" \|
	~~The BluePippin has~~ four different percentages of agarose gels that can be used for different size selections. A stock of most types of cartridges are kept in the BioMicro Center. For long fragments using the pulse field, there are multiple settings on the BluePippin, each optimized for a different size range.		Both Pippins can utilize four different percentages of agarose gels that can be used for different size selections. A stock of most types of cartridges are kept in the BioMicro Center. For long fragments using the pulse field, there are multiple settings on the BluePippin, each optimized for a different size range.
	\|		\|
	{\| border="1" align="right" style="text-align: center"		{\| border="1" align="right" style="text-align: center"

imported>Alexei at 17:37, 1 June 2020

2020-06-01T17:37:59Z

New page

{{BioMicroCenter}}

The BioMicroCenter utilizes the Sage Scientific Blue Pippin ;http://www.sagescience.com/wp-content/uploads/2016/09/BluePippin-Operations-Manual-460013-Rev-D.pdf] for any gel based sizing. The instrument can be run as a walk-up or as an assisted service as part of Illumina, PacBio, or Oxford Nanopore library prep. 

== Preparative Gel Electrophoresis ==
{| align=left
|- style="vertical-align: top;"
|style="width: 300px;"|
{| class="wikitable" border=1
! Instrument || Pippin/BluePippin
|-
| TYPE || WALKUP or ASSISTED assisted service available as part of library preparation
|-
| UNIT || Per run for walkup Per lane for assisted
|-
| FORMAT || 5 lane gel from 0.75% agarose to 3% agarose
|-
| INPUT || >1ug of DNA in low salt buffer
|-
| EXPECTED YIELD || 10-50% of material in the selected size range Narrower ranges result in lower yield.
|-
| WALKUP SIGNUP || [https://mit.ilabsolutions.com/equipment/show/262716/?tab=schedule ilabs calendar]
|-
| NEW USERS || New users should request training by emailing biomicro@mit.edu
|-
| DONATED BY || Department of Biology
|}
|
The BluePippin (Sage Science) is an automated preparative gel electrophoresis system that simplifies size fractionation of DNA. Runs of the five lane gels take approximately 1 hour to overnight depending on the fragment length and can either use internal/internal lane markers for calibration or ethidium bromide to detect passing bands. The BluePippin can separate bands between 50bp and 50kb. We have done some testing of the BluePippin system and the results are [[BioMicroCenter:PippinPrep_Testing|'''HERE''']]. 

Critically, the system does require large amounts of input. Yields of <50% are typical '''within the desired range''' so multiple ug of input are strongly recommended before using the Pippin systems. Samples with lower amounts of input should consider alternative approaches including SPRIbead cuts. 

The system is based on precast gel cartridges. These gels narrow as they run, concentrating the sample from the initial 40ul (30ul sample + 10ul loading buffer) loaded in the wells. The BluePippin uses an optical detector to measure the rate of passage of bands based on high speed fluorescent markers or by directly detecting the DNA using ethidium bromide. Once the desired band passes the detector, the valve on each lane switches from a waste reservoir to an elution chamber. Once the band is passed, the valve turns back to waste and the desired band can be removed from the elution chamber with a pippette. Samples can be cleaned up with either SPRI beads or using an ethanol precipitation.
|
[[Image:BMC_Pippin.jpg|thumb|right|250px|BluePippin]]
[[Image:BMC_pippinGel.png|thumb|right|250px|BluePippin Gel]]

|- style="vertical-align: top;"
| colspan="2" |
The BluePippin has four different percentages of agarose gels that can be used for different size selections. A stock of most types of cartridges are kept in the BioMicro Center. For long fragments using the pulse field, there are multiple settings on the BluePippin, each optimized for a different size range.
|
{| border="1" align="right" style="text-align: center"
|-
!GEL
!SIZE RANGE
!NOTES
|-
|3.0%
|50-200bp
|Used for small RNA libraries
|-
|2.0%
|100-600bp
|
|-
|1.5%
|0.3-1.5kb
|Used for RNAseq libraries
|-
|0.75%
|1.0-50kb
|Uses pulse field.
|}
|}