Recombinant and in vitro reconstitution methods for producing lasso peptides are provided. Methods of screening lasso peptides are also provided.

The present invention provides a screening method of negative regulatory factors of a Streptomyces biosynthesis gene cluster, the method including: constructing a reporter system in a Streptomyces cell, which is mediated by a promoter of a self-owned target gene of the Streptomyces cell, and then randomly mutating Streptomyces with the reporter system by using a random mutation system constructed based on a transposon Himar1; intensively screening Streptomyces strains that have been subjected to random mutation to obtain a Streptomyces strain with high expression of the target gene; performing phage packaging on a genome of the Streptomyces strain with high expression of the target gene and screening out a cosmid with a random insert; and determining the position of the random insert in the genome of the Streptomyces strain with high expression of the target gene by sequencing DNAs of the cosmid.

The present invention provides a screening method of negative regulatory factors of a Streptomyces biosynthesis gene cluster, the method including: constructing a reporter system in a Streptomyces cell, which is mediated by a promoter of a self-owned target gene of the Streptomyces cell, and then randomly mutating Streptomyces with the reporter system by using a random mutation system constructed based on a transposon Himar1; intensively screening Streptomyces strains that have been subjected to random mutation to obtain a Streptomyces strain with high expression of the target gene; performing phage packaging on a genome of the Streptomyces strain with high expression of the target gene and screening out a cosmid with a random insert; and determining the position of the random insert in the genome of the Streptomyces strain with high expression of the target gene by sequencing DNAs of the cosmid.

Disclosed is a synthetic gene cluster for producing gadusol derivatives, expression vectors and host cells containing the same, methods of producing gadusol derivatives, and compositions thereof. In an example, the synthetic gene cluster includes a valA nucleotide sequence capable of expressing ValA protein; a nucleotide sequence capable of expressing methyltransferase/oxidoreductase (MT-Ox) protein; a mysC nucleotide sequence capable of expressing a MysC protein; and a mysD nucleotide sequence capable of expressing a MysD protein. In this way, gadusol derivatives can be produced in amounts sufficient for use in a variety of applications.

The present invention relates to a DoxA protein mutant having an amino acid sequence set forth in SEQ ID No. 16, and coding gene thereof. The protein mutant or the coding gene thereof can be used for producing epirubicin. The present invention further relates to a Streptomyces capable of efficiently expressing epirubicin, which is constructed by replacing the dnmV gene of a starting Streptomyces in situ with the avrE gene and mutating the doxA gene of the starting Streptomyces into a gene encoding the protein set forth in SEQ ID No. 16. The fermentation broth of this Streptomyces has an epirubicin potency of up to 102.0 g/ml.

The present invention relates to a DoxA protein mutant having an amino acid sequence set forth in SEQ ID No. 16, and coding gene thereof. The protein mutant or the coding gene thereof can be used for producing epirubicin. The present invention further relates to a Streptomyces capable of efficiently expressing epirubicin, which is constructed by replacing the dnmV gene of a starting Streptomyces in situ with the avrE gene and mutating the doxA gene of the starting Streptomyces into a gene encoding the protein set forth in SEQ ID No. 16. The fermentation broth of this Streptomyces has an epirubicin potency of up to 102.0 g/ml.

The present invention relates to a Streptomyces mutant strain in which a -agarase DagA enzyme is over-expressed and a method for developing the strain. In addition, the present invention relates to a method for producing neoagarohexaose or neoagarotetraose in vivo by using the Streptomyces mutant strain.

The present invention relates to a Streptomyces mutant strain in which a -agarase DagA enzyme is over-expressed and a method for developing the strain. In addition, the present invention relates to a method for producing neoagarohexaose or neoagarotetraose in vivo by using the Streptomyces mutant strain.

This disclosure describes compositions and methods for enhanced production of enduracidin in genetically engineered strains of Streptomyces fungicidicus. In particular, the present disclosure describes the genetic manipulation of regulatory genes orf24 and orf18 associated with the enduracidin (enramycin) biosynthesis gene cluster from Streptomyces fungicidicus to generate vector constructs and recombinant strains producing greater yields of enduracidin.

The present disclosure includes refactored thaxtomin biosynthetic gene clusters including thaxtomin modules including one or more thaxtomin genes such that the expression of the refactored thaxtomin biosynthetic gene cluster produces at least one thaxtomin compound in the absence of thaxtomin-inducing conditions. Also included are genetically engineered Streptomyces bacterium from a non-pathogenic Streptomyces strain comprising an exogenous, refactored thaxtomin biosynthetic gene cluster of the present disclosure, such that the expression of the refactored thaxtomin biosynthetic gene cluster provides the genetically engineered Streptomyces bacterium with the ability to produce at least one thaxtomin compound in the absence of thaxtomin-inducing conditions. The present disclosure also includes methods of producing thaxtomin compounds, analogs, or intermediate with the refactored thaxtomin biosynthetic gene clusters and genetically engineered bacteria of the present disclosure.