CN105950653A - Universal plasmid of bombyx mori middle silkgland bioreactor as well as construction method and application of universal plasmid - Google Patents
Universal plasmid of bombyx mori middle silkgland bioreactor as well as construction method and application of universal plasmid Download PDFInfo
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Abstract
The invention discloses a universal plasmid of a bombyx mori middle silkgland bioreactor as well as a construction method and an application of the universal plasmid. A sericin 1 gene promoter, a His 6 sequence, a DDDDK sequence, light-chain fibroin signal peptide, a light-chain fibroin gene 3' end sequence, a bombyx mori actin gene A3 promoter, an EGFP gene and an SV40 sequence are obtained, and then the plasmid is constructed; a first expression cassette includes the A3 promoter, the EGFP gene and the SV40 sequence; and a second expression cassette includes the sericin 1 gene promoter, the light-chain fibroin signal peptide, the His 6 sequence, the DDDDK and the light-chain fibroin gene 3' end sequence, as well as two restriction sites, namely ApaI and NheI. The plasmid disclosed by the invention can reduce workload of constructing the donor plasmid and can improve the working efficiency of the bombyx mori middle silkgland bioreactor.
Description
Technical field
The present invention relates to a kind of plasmid and construction method thereof and application, especially related in the middle part of a kind of silkworm
The universal plasmid of sericterium bioreactor and construction method thereof and application.
Background technology
PiggyBac transposon is initially the base from cabbage looper (Trichoplusia ni) TN-368 cell strain
Because of isolated in group, it it is the DNA transposon that the transposition activity having now been found that is the highest.PiggyBac turns
Base system is a kind of non-virus carrier, and transposition efficiency is higher.Compared with sleeping beauty, piggyBac
Bearer capabilities is relatively big, portability 18kb, it is possible to achieve polygenic coexpression, and swivel base fragment cut after not
Can leave trace (footprint) in position, genome is accurately repaired, at reversible base after can realizing excision
The application of cause has important function.
The more than ten years have been carried out in the research of the transgenic bombyx mori sericterium bioreactor of piggyBac transposon mediation,
The scientist of countries in the world is devoted to the expression of exogenous gene, has built up with fibroin albumen light chain promoter
(Fib-L Promoter), fibroin albumen heavy chain promoter (Fib-H Promoter) and sericin 1 start
Son (Ser1Promoter) expresses the transgenic bombyx mori sericterium bioreactor of foreign protein.
Owing to piggyBac carrier majority contains two expression cassettes, there is more element and identical enzyme action position
Point, so, when expressing different exogenous genes, need from the beginning to build plasmid every time, assemble promoter, letter
Number structure such as peptide, exogenous gene, polyA, wastes time and energy, and work efficiency is low.
Summary of the invention
In order to solve problem present in background technology, it is an object of the invention to propose in the middle part of a kind of silkworm
The universal plasmid of sericterium bioreactor and construction method thereof and application, specifically utilize routine to build plasmid
Biological technique method builds one and does not contains genes of interest, but has the universal plasmid of other all required elements.
In order to achieve the above object, the step of the technical solution used in the present invention is as follows:
One, a kind of silkworm universal plasmid of middle division of silkgland bioreactor:
Described plasmid is pBac [A3-EGFP-SV40]-[Ser1promoter-FLSP-His 6-DDDDK-FL
PA], it is based on piggyBac transposon and with Amp resistant gene, including piggyBac transposon
Two swivel base arm PBL and PBR and two swivel base arms between two functional expression frames.
One functional expression frame is the Green Fluorescent Protein Gene Expression frame that A3 promoter starts, i.e. A3
Promoter EGFP-SV40, another functional expression frame be comprise domestic silkworm silk glue protein 1 gene promoter,
Fibroin albumen light chain gene signal peptide, hexahistine, enterokinase cleavage site and silk fibroin protein light chain
The expression cassette of gene 3 ' end, i.e. Ser1Promoter-Fibroin L chain signal peptide-His 6-
DDDDK-Fibroin L chain PolyA。
Containing two restriction enzyme sites of narrow spectrum ApaI and NheI between described DDDDK-FLPA, two
Individual restriction enzyme site is formed with the plasmid of function after plasmid specificity is cut connection exogenous gene.
Two, the construction method of a kind of silkworm universal plasmid of middle division of silkgland bioreactor, the step of method is such as
Under:
1) [sericin 1 gene promoter 575-Renilla luciferase is comprised by molecular biology method structure
Gene-SV40]-[sericin 1 gene promoter 4023-Fluc gene-SV40]-[A3 gene
Promoter-green fluorescence protein gene-SV40] ([Ser 575-Rluc-SV40]-[Ser
4023-Fluc-SV40]-[A3-GFP-SV40]) plasmid piggy-14413, with plasmid piggy-14413 be
Template, the base sequence of plasmid piggy-14413 such as SEQ ID NO.1, with such as SEQ ID NO.2
GgtaccGTTGGCGGTCTTTGGATCG sequence 1 and such as SEQ ID NO.3
GaattccttaagGCACACACACTACATACCATGTATTTG sequence 2 is primer, uses PCR
Amplification obtains sericin 1 gene promoter of a length of 583bp of fragment, its base sequence such as SEQ ID NO.4；
2) fib-L gene promoter-fibroin light chain protein signal is comprised with EcoR I and Kpn I double digestion
Peptide-hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence (FL
692-FLSP-His 6-DDDDK-FLPA) and the plasmid piggy-4086 of Amp resistant gene, plasmid
Its base sequence of piggy-4086 such as SEQ ID NO.5, obtain a length of 3378bp comprises fibroin light chain egg
White signal peptide-hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence
The fragment of (FLSP-His 6-DDDDK-FLPA)；
3) Connection Step 1) sericin 1 gene promoter that obtains and step 2) obtain comprise fibroin
Light chain protein signal peptide-hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence
Fragment, obtains comprising sericin 1 gene promoter-fibroin light chain protein signal peptide-hexahistine-intestinal and swashs
Enzyme restriction enzyme site sequence-silk fibroin protein light chain 3 ' sequence (Ser1 promoter-FLSP-His
6-DDDDK-FLPA) and the plasmid piggy-3969 of Amp resistant gene；
4) by AflII and BglII double digestion step 3) the plasmid piggy-3969 that obtains, it is thus achieved that a length of 1322bp
Comprise sericin 1 gene promoter-fibroin light chain protein signal peptide-hexahistine-enterokinase enzyme action position
The fragment of point sequence-silk fibroin protein light chain 3 ' sequence (Ser-FLSP-His 6-DDDDK-FLPA)；
5) A3 gene promoter-green fluorescence protein gene-SV40 is comprised with AflII and BglII double digestion
And the plasmid piggy-6212 of Amp resistant gene, the base of plasmid piggy-6212 (A3-GFP-SV40)
Sequence such as SEQ ID NO.6, obtain a length of 5870bp comprises A3 gene promoter-green fluorescent protein
The fragment of gene-SV40 (A3-GFP-SV40)；
6) Connection Step 4) obtain comprise sericin 1 gene promoter-fibroin light chain protein signal peptide-
The fragment of hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence and step 5)
The fragment comprising A3 gene promoter-green fluorescence protein gene-SV40 obtained, is comprised [A3 gene
Promoter-green fluorescence protein gene-SV40]-[sericin 1 gene promoter-fibroin light chain protein signal peptide-
Hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence
The plasmid of ([A3-EGFP-SV40]-[Ser1 promoter--FLSP-His-DDDDK-FLPA])
Piggy-7192, as plasmid of the present invention, is the silkworm universal plasmid of middle division of silkgland bioreactor,
Its base sequence such as SEQ ID NO.7.
Three, described plasmid is for the application of the donor plasmid of the various exogenous gene of construction expression.
The present invention first designs primer, obtains the sequence of sericin 1 gene promoter by the method for PCR；
The most respectively by the corresponding fragment of digestion with restriction enzyme, after electrophoresis obtains purpose band, cut glue and reclaim, use
Corresponding fragment is connected by T4 ligase, makes required element (Ser1Promoter-Fibroin L chain
Signal peptide-His6-DDDDK-Fibroin L chain PolyA) it is sequentially connected with；Then by piggy-6212
Marker gene in plasmid and said elements are incorporated into same plasmid, obtain universal plasmid, final such as Fig. 1
Shown in.
The invention has the beneficial effects as follows:
The plasmid of the present invention only need to can just will through a step enzyme action coupled reaction for any one exogenous gene
Any exogenous gene imports plasmid, completes the structure of expression alien gene plasmid.
Accompanying drawing explanation
Fig. 1 is the composition schematic diagram of the universal plasmid of Ser1 of the present invention.
Detailed description of the invention
The invention will be further described with embodiment below in conjunction with the accompanying drawings.
Embodiments of the invention are as follows:
(1) design primer, with piggy-14413 plasmid (SEQ ID NO.1) as template, it is thus achieved that sericin gene opens
Mover (SEQ ID NO.4).And add the restriction enzyme site of EcoR I at promoter 5 ' end, add at 3 ' ends
The restriction enzyme site of KpnI.By EcoR I and KpnI double digestion PCR primer, obtain the fragment of 593bp,
I.e. promoter, glue reclaims；With EcoR I and KpnI double digestion piggy-4086 plasmid (SEQ ID NO.5),
Obtaining the fragment of 3378bp, glue reclaims；Connect two purpose fragments, obtain piggy-3969 plasmid.
(2) with AflII and BglII, ScaI enzyme action piggy-3969 plasmid, the fragment of a length of 1322bp is obtained；
With AflII and BglII double digestion piggy-6212 plasmid (SEQ ID NO.6), obtain a length of 5870bp
Fragment, connect above-mentioned two purpose fragments, obtain piggy-7192 plasmid (SEQ ID NO.7), be house
The universal plasmid of silkworm middle division of silkgland bioreactor.
(3) with ApaI and NheI double digestion piggy-7192 plasmid, comprising of a length of 7190bp is obtained
[A3-EGFP-SV40]-[Ser1 promoter-FLSP-His 6-DDDDK-FLPA and Amp resistant gene
Gene order.Upstream extremity is glued with NheI enzyme action with ApaI enzyme action cohesive end sequence, downstream
The T4 ligase gene of property end sequence is connected with above-mentioned fragment, is built into silkworm middle division of silkgland bioreactor
Expressing the plasmid piggy-8716 of T4 ligase gene, the expression cassette sequence that this plasmid comprises is
[A3-EGFP-SV40]-[Ser1promoter-FLSP-His 6-DDDDK-T4Ligase-FLPA]。
The above results illustrates, utilizes the silkworm middle division of silkgland universal plasmid of bioreactor double-promoter
Piggy-7912, only the step silkworm middle division of silkgland that just can obtain being in expresses the plasmid of T4 ligase gene.
Illustrate to utilize the silkworm middle division of silkgland bioreactor universal plasmid construction transgenic bombyx mori of the present invention
Donor plasmid, operation sequence is simple, and work efficiency is high.
Claims (5)
1. the silkworm universal plasmid of middle division of silkgland bioreactor, it is characterised in that: described plasmid is
PBac [A3-EGFP-SV40]-[Ser1promoter-FLSP-His 6-DDDDK-FLPA], is with piggyBac
Based on transposon and with Amp resistant gene, including two swivel base arm PBL of piggyBac transposon
And two functional expression frames between PBR and two swivel base arms.
A kind of silkworm universal plasmid of middle division of silkgland bioreactor the most according to claim 1, it is special
Levy and be: a functional expression frame is the Green Fluorescent Protein Gene Expression frame that A3 promoter starts, i.e. A3
Promoter EGFP-SV40, another functional expression frame be comprise domestic silkworm silk glue protein 1 gene promoter,
Fibroin albumen light chain gene signal peptide, hexahistine, enterokinase cleavage site and silk fibroin protein light chain
The expression cassette of gene 3 ' end, i.e. Ser1 Promoter-Fibroin L chain signal peptide-His 6-
DDDDK-Fibroin L chain PolyA。
A kind of silkworm universal plasmid of middle division of silkgland bioreactor the most according to claim 1, it is special
Levy and be: between described DDDDK-FLPA, contain two restriction enzyme sites of narrow spectrum ApaI and NheI,
Two restriction enzyme sites are formed with the plasmid of function after plasmid specificity is cut connection exogenous gene.
4. the construction method of the silkworm universal plasmid of middle division of silkgland bioreactor, it is characterised in that method
Step as follows:
1) [sericin 1 gene promoter 575-Renilla luciferase is comprised by molecular biology method structure
Gene-SV40]-[sericin 1 gene promoter 4023-Fluc gene-SV40]-[A3 gene
Promoter-green fluorescence protein gene-SV40] plasmid piggy-14413, with plasmid piggy-14413 as mould
Plate, with the ggtaccGTTGGCGGTCTTTGGATCG sequence 1 such as SEQ ID NO.2 with such as SEQ ID
The gaattccttaagGCACACACACTACATACCATGTATTTG sequence 2 of NO.3 is primer,
Obtain sericin 1 gene promoter of a length of 583bp of fragment, its base sequence such as SEQ ID NO.4；
2) fib-L gene promoter-fibroin light chain protein signal is comprised with EcoR I and Kpn I double digestion
Peptide-hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence (FL
692-FLSP-His 6-DDDDK-FLPA) and the plasmid piggy-4086 of Amp resistant gene, plasmid
Its base sequence of piggy-4086 such as SEQ ID NO.5, obtain a length of 3378bp comprises fibroin light chain egg
The fragment of white signal peptide-hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence；
3) Connection Step 1) sericin 1 gene promoter that obtains and step 2) obtain comprise fibroin
Light chain protein signal peptide-hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence
Fragment, obtains comprising sericin 1 gene promoter-fibroin light chain protein signal peptide-hexahistine-intestinal and swashs
Enzyme restriction enzyme site sequence-silk fibroin protein light chain 3 ' sequence and the plasmid piggy-3969 of Amp resistant gene；
4) by AflII and BglII double digestion step 3) the plasmid piggy-3969 that obtains, it is thus achieved that a length of 1322bp
Comprise sericin 1 gene promoter-fibroin light chain protein signal peptide-hexahistine-enterokinase enzyme action position
Point sequence-silk fibroin protein light chain 3 ' sequence (Ser1 promoter-FLSP-His 6-DDDDK-FLPA)
Fragment；
5) A3 gene promoter-green fluorescence protein gene-SV40 is comprised with AflII and BglII double digestion
And the plasmid piggy-6212 of Amp resistant gene, the base of plasmid piggy-6212 (A3-GFP-SV40)
Sequence such as SEQ ID NO.6, obtain a length of 5870bp comprises A3 gene promoter-green fluorescent protein
The fragment of gene-SV40；
6) Connection Step 4) obtain comprise sericin 1 gene promoter-fibroin light chain protein signal peptide-
The fragment of hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence and step 5)
The fragment comprising A3 gene promoter-green fluorescence protein gene-SV40 obtained, is comprised [A3 gene
Promoter-green fluorescence protein gene-SV40]-[sericin 1 gene promoter-fibroin light chain protein signal peptide-
The plasmid piggy-7192 of hexahistine-enterokinase cleavage site sequence-silk fibroin protein light chain 3 ' sequence
Its base sequence such as SEQ ID NO.7.
5. the application of the silkworm universal plasmid of middle division of silkgland bioreactor, it is characterised in that: right is wanted
Ask the plasmid described in 1～3 for the donor plasmid of the various exogenous gene of construction expression.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111793644A (en) * | 2020-07-17 | 2020-10-20 | 西南大学 | Silkworm fibroin heavy chain expression system and preparation method and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102226202A (en) * | 2011-05-13 | 2011-10-26 | 浙江大学 | Method for synthetizing secretion lysozyme by middle silkgland cell of silkworm |
| CN104593413A (en) * | 2014-12-31 | 2015-05-06 | 浙江大学 | Method for synthesizing secreted human serum albumin employing bombyx mori posterior silk gland |
| CN104846011A (en) * | 2015-03-18 | 2015-08-19 | 浙江大学 | Method for synthesizing royal jelly main protein 1 by using bombyx mori posterior silkgland |
-
2016
- 2016-05-04 CN CN201610287168.4A patent/CN105950653A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102226202A (en) * | 2011-05-13 | 2011-10-26 | 浙江大学 | Method for synthetizing secretion lysozyme by middle silkgland cell of silkworm |
| CN104593413A (en) * | 2014-12-31 | 2015-05-06 | 浙江大学 | Method for synthesizing secreted human serum albumin employing bombyx mori posterior silk gland |
| CN104846011A (en) * | 2015-03-18 | 2015-08-19 | 浙江大学 | Method for synthesizing royal jelly main protein 1 by using bombyx mori posterior silkgland |
Non-Patent Citations (1)
| Title |
|---|
| 钟伯雄等: "piggyBac转座子介导的转基因家蚕丝腺生物反应器研究进展", 《中国农业科学》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111793644A (en) * | 2020-07-17 | 2020-10-20 | 西南大学 | Silkworm fibroin heavy chain expression system and preparation method and application thereof |
| CN111793644B (en) * | 2020-07-17 | 2023-10-20 | 西南大学 | Home silk fibroin heavy chain expression system and preparation method and application thereof |
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