Uniclone One Step Seamless Cloning Kit Ver. 2(SC613)

English 简体中文

Welcome to Genesand Biotech Co.,Ltd

0086010-400-688-7890

To help life science research,

let more research results benefit mankind

To help life science research,

let more research results benefit mankind

PRODUCTS

HOME >> Cloning >> One Step Rapid Cloning >> Uniclone One Step Seamless Cloning Kit Ver. 2(SC613)

Uniclone One Step Seamless Cloning Kit Ver. 2(SC613)

List price：＄ 230.00

Price：＄ 230.00

Amount：

50 rxns

Cat. No.：

SC613

Quantity：

Buy now

Add to cart

Product Description

Highlights

1) Efficient: it has higher efficiency in recombining multiple fragments.

2) Quick: the reorganization can be completed within 5~60 minutes.

3) Simple: there is no need for fragment enzyme digestionit and not limited by the restriction of enzyme digestion sites;.

4) Seamless: no additional sequences are introduced.

Introduction

Uniclone One Step Seamless Cloning Kit Ver. 2 is a simple, fast and efficient seamless cloning kit developed based on the principle of homologous recombination. It can recombine any DNA fragment containing the overlapping region of the end of the vector onto the linearized vector, which is not limited by the restriction of enzyme digestion sites, and has the low vector self ligation background. This method requires adding a 15-25 nt sequence at the end of the linearized vector to the 5’end of the forward / reverse PCR primer of the inserted fragment, so that the fragment and the vector can be recombined through the action of recombinant enzyme because of overlapping region.

Composition

Components

Amount(50 rxns)

2×Uniclone Seamless Cloning Mix Ver. 2

250 μL

pUC19 Control Plasmid,Linearized(Ampr,40 ng/μl) *

5 μL

Control Fragment(500 bp,20 ng/μl)**

5 μL

* And ** respectively represent control linearized vector and control linearized fragment, these two components are connected with each other, which can eliminate the influencing factors of the experiment.

Related Products

Type	Composition	Cat. No.	Amount	Introduction
Nucleic Acid Isolation	DNA Gel Recovery Kit	GE706	200 rxns	One box of dual-purpose, can effectively recover 80 bp ~10 kb double-stranded DNA fragments, can also directly recover PCR stock solution, gel without weighing, easy to operate, recovery rate of up to 85%
High-Fidelity PCR	1.1×S4 Fidelity PCR Mix(dye+)	SF212	5×1.125 mL	It is a 1.1 × ready-to-use mixture for efficient PCR amplification, which has high fidelity and extremely fast extension speed.
Plasmid DNA Extraction	High Purity Plasmid DNA Mini-Extraction Kit	PE707	200 rxns	Improved SDS alkali lysis method, high-purity plasmid can be extracted in 30 min

Q1: No or Few Colonies Obtained from Transformation?

A1:

1) Low transformation efficiency. Bacteria were not competent. Check transformation efficiency by the control plasmid. You should obtain >1 x 108 cfu/μg; otherwise use fresh competent cells.

2) Transformed with too much/little reaction, see above protocol for details.

3) The PCR product was not purified or spent too much time under UV irradiation . It is recommended to be operated at 365 nm long wave UV.

4) Linearized vector and insert fragment are not pure and inhibit reaction. The experiment proves that EDTA and guanidine in products which were not fully purified can significantly reduce the recombinant efficiency. It should be purified again.

Q2: Large Numbers of Colonies Contained No Insert?

A2:

1) Incomplete linearization of your vector. It is important to remove any uncut vector prior to use in Sosoo reaction. If necessary, recut your vector and gel purify.

2) Contamination of Sosoo reaction by plasmid with same antibiotic resistance. If your insert was amplified from a plasmid, closed circular DNA may have carried through purification and contaminated the cloning reaction. Firstly, you must ensure the removal of any plasmid contamination, we recommend linearizing the template DNA before performing PCR. Secondly, If you spin-column purify your insert, treating the PCR product with DpnI before purification will help to remove contaminating template DNA.

Q3: Clones Contained Incorrect Insert?

A3:

1) Your PCR product contained non-specific sequences. If your PCR product is not a single distinct band, then it may be necessary to optimize your PCR protocol or gel purify the PCR product to ensure cloning of the correct insert.

2) Polluted by plasmid with same antibiotic resistance. If your linearized vector is not obtained by empty vector but by the vector that had been inserted into the other fragments , it may result in the some background and make some clone contains incorrect insertion fragment. Improve the efficiency of enzyme, treating the PCR product with DpnI before purification, using adbanced linearized vector as PCR template all can effectively avoid such situation happen.